Q.1 What is the primary advantage of Sanger sequencing compared to early next-generation sequencing methods?
It can sequence millions of reads in a single run
It produces very long reads with low error rates
It requires a small amount of DNA for library preparation
It allows sequencing of a single DNA molecule without amplification
Explanation - Sanger sequencing typically yields read lengths of 800–1000 base pairs and has a very low error rate (~0.001), which was a key advantage before high-throughput methods were developed.
Correct answer is: It produces very long reads with low error rates
Q.2 Which of the following is NOT a step in preparing a DNA library for Illumina sequencing?
Fragmentation of DNA
End-repair and A-tailing
Size selection
Amplification using PCR
Explanation - Illumina library prep normally avoids PCR amplification to reduce bias; instead it uses enzymatic steps like end‑repair and A‑tailing, followed by adapter ligation and size selection.
Correct answer is: Amplification using PCR
Q.3 What is a typical read length range for Illumina short‑read sequencers?
50–150 base pairs
300–500 base pairs
1–2 kilobases
10–20 kilobases
Explanation - Standard Illumina platforms (e.g., HiSeq, NovaSeq) produce reads typically 50–150 bp long, which is suitable for many applications such as RNA‑seq and variant calling.
Correct answer is: 50–150 base pairs
Q.4 Which sequencing technology uses a polymerase that incorporates fluorescently labeled nucleotides and reads the signal in a flow cell?
Nanopore sequencing
Ion Torrent sequencing
Illumina sequencing
PacBio sequencing
Explanation - Illumina’s sequencing‑by‑synthesis platform uses reversible dye terminators to detect the incorporation of each nucleotide in a flow cell.
Correct answer is: Illumina sequencing
Q.5 The main source of error in Nanopore sequencing is due to:
Insertion–deletion errors in homopolymer stretches
Random base substitution errors
Base‑calling algorithm inaccuracies
Loss of signal during the translocation step
Explanation - Nanopore sequencing generates electrical signals that are translated into bases by complex algorithms; inaccuracies in these algorithms cause high error rates, especially in homopolymers.
Correct answer is: Base‑calling algorithm inaccuracies
Q.6 Which of the following best describes the concept of 'coverage' in genome sequencing?
The number of different DNA molecules sequenced in a run
The total number of bases sequenced per million reads
The average number of times a single base in the genome is read
The length of the longest read obtained
Explanation - Coverage, or depth, refers to how many times, on average, each base in the target genome is sequenced, which influences confidence in variant calls.
Correct answer is: The average number of times a single base in the genome is read
Q.7 In Illumina sequencing, the 'bridge amplification' step occurs on which surface?
The liquid‑phase in the sequencing tube
The flow‑cell surface
The polymerase‑immobilized beads
The magnetic bead surface
Explanation - Bridge amplification forms clusters of identical DNA fragments directly on the flow‑cell surface by hybridizing and amplifying the fragments in situ.
Correct answer is: The flow‑cell surface
Q.8 Which sequencing platform is known for producing the longest read lengths, often exceeding 10 kb?
Illumina HiSeq
Ion Torrent Proton
PacBio Sequel II
Sanger Sequencer
Explanation - PacBio’s SMRT sequencing can routinely produce reads over 10 kb, and the Sequel II system can generate even longer reads with improved accuracy.
Correct answer is: PacBio Sequel II
Q.9 Which of the following best explains a 'homopolymer' in DNA sequencing?
A stretch of identical bases
A sequence with alternating purines and pyrimidines
A region with high GC content
A region with a high density of SNPs
Explanation - A homopolymer consists of repeated identical nucleotides, such as AAAAA, which can challenge base‑calling in platforms like Ion Torrent and Nanopore.
Correct answer is: A stretch of identical bases
Q.10 What is the key advantage of using a PCR‑free library preparation for Illumina sequencing?
Reduced sequencing cost
Elimination of bias introduced by PCR amplification
Increased read length
Simplified sample handling
Explanation - PCR amplification can introduce bias against certain sequences; a PCR‑free protocol preserves the original representation of the DNA in the sample.
Correct answer is: Elimination of bias introduced by PCR amplification
Q.11 Which sequencing technology detects hydrogen ions released during DNA synthesis?
Ion Torrent PGM
Illumina NovaSeq
Nanopore MinION
PacBio Sequel
Explanation - Ion Torrent measures pH changes (hydrogen ions) as nucleotides are incorporated, converting them into electrical signals.
Correct answer is: Ion Torrent PGM
Q.12 Why is 'paired‑end sequencing' useful in genome assembly?
It doubles the read length
It provides information about the orientation and distance between two reads from the same DNA fragment
It reduces the cost per base
It eliminates PCR amplification bias
Explanation - Paired‑end reads, sequenced from both ends of a fragment, help to resolve repeats and structural variations by indicating the expected distance and orientation between reads.
Correct answer is: It provides information about the orientation and distance between two reads from the same DNA fragment
Q.13 Which base‑calling algorithm is most commonly used for Illumina sequencing data?
GATK
bcl2fastq
Nanopolish
SignalAlign
Explanation - bcl2fastq is Illumina’s software for converting BCL files generated by the sequencer into FASTQ files and performing basic base‑calling.
Correct answer is: bcl2fastq
Q.14 The 'error profile' of a sequencing platform refers to:
The distribution of errors across the read length and sequence context
The cost per base of sequencing
The maximum read length achievable
The throughput of the sequencer
Explanation - An error profile describes where and how often errors occur, such as higher error rates near the ends of reads or within certain motifs.
Correct answer is: The distribution of errors across the read length and sequence context
Q.15 Which of the following is a major challenge when using Nanopore sequencing for de‑novo genome assembly?
Very short read lengths
High substitution error rates
Very low throughput
Inability to sequence GC‑rich regions
Explanation - Nanopore sequencing historically has higher substitution error rates compared to short‑read platforms, making accurate assembly of complex genomes more difficult without error correction.
Correct answer is: High substitution error rates
Q.16 What does the 'Q-score' represent in sequencing quality metrics?
The number of bases in a read
The probability of a base being called incorrectly
The speed of sequencing
The read length distribution
Explanation - A Q-score of 30 corresponds to a 1 in 1000 chance of a wrong base call, calculated as Q = –10 log10(p).
Correct answer is: The probability of a base being called incorrectly
Q.17 Which sequencing method uses a template that is circularized for continuous sequencing cycles?
Illumina sequencing
Nanopore sequencing
PacBio SMRT sequencing
Ion Torrent sequencing
Explanation - PacBio’s SMRTbell library forms a circular template, allowing the polymerase to read the same molecule repeatedly, improving accuracy via circular consensus sequencing.
Correct answer is: PacBio SMRT sequencing
Q.18 The term 'throughput' in sequencing refers to:
The length of the longest read
The number of reads generated in a sequencing run
The cost per base
The time required to finish sequencing
Explanation - Throughput measures how many base pairs or reads are produced during a sequencing run, a key factor for large projects.
Correct answer is: The number of reads generated in a sequencing run
Q.19 Which of the following is a typical application of Oxford Nanopore sequencing?
Whole genome sequencing of bacterial isolates in a clinical setting
High‑throughput small‑variant discovery in human genomes
Targeted sequencing of exomes for diagnostic purposes
RNA‑seq for gene expression analysis
Explanation - Nanopore’s portability and long reads make it well suited for rapid, field‑based sequencing of pathogens in clinical microbiology.
Correct answer is: Whole genome sequencing of bacterial isolates in a clinical setting
Q.20 In the context of DNA sequencing, what is 'adapter contamination'?
Residual sequencing primers in the final library
Short sequences added to DNA ends that are not removed before analysis
Unwanted bacterial DNA in the sample
PCR duplicates that mimic genuine reads
Explanation - Adapters are ligated during library prep; if not trimmed, they remain in the reads and can interfere with alignment and variant calling.
Correct answer is: Short sequences added to DNA ends that are not removed before analysis
Q.21 Which sequencing platform was the first to be marketed as a 'next‑generation' sequencer?
Illumina Solexa
Pacific Biosciences RS
Ion Torrent PGM
Roche 454
Explanation - Illumina’s Solexa platform, introduced in 2006, pioneered sequencing‑by‑synthesis and became the dominant NGS platform.
Correct answer is: Illumina Solexa
Q.22 Which of the following best describes 'circular consensus sequencing' (CCS) in PacBio?
Sequencing a circular DNA template multiple times to improve accuracy
Sequencing both strands of a duplex DNA molecule simultaneously
Using a circular flow cell to increase throughput
Amplifying circular plasmids before sequencing
Explanation - CCS generates highly accurate reads by repeatedly sequencing the same circular template, averaging out random errors.
Correct answer is: Sequencing a circular DNA template multiple times to improve accuracy
Q.23 Which of the following is a major disadvantage of Ion Torrent sequencing?
Inability to detect indels
Very low read accuracy
High sequencing cost
Poor performance on homopolymers
Explanation - Ion Torrent’s pH‑based detection struggles with accurately counting the number of bases in homopolymer tracts, leading to indel errors.
Correct answer is: Poor performance on homopolymers
Q.24 During Illumina sequencing, the 'phasing' error occurs when:
Some DNA strands incorporate nucleotides out of sync with the cluster
The flow cell temperature fluctuates
The polymerase stalls on a template
The fluorescent signal is too weak
Explanation - Phasing refers to a mismatch between the timing of nucleotide incorporation across the cluster, causing signal decay and errors.
Correct answer is: Some DNA strands incorporate nucleotides out of sync with the cluster
Q.25 Which sequencing platform can be directly coupled to a portable sequencer for field work?
Illumina MiSeq
Pacific Biosciences RS II
Oxford Nanopore MinION
Roche 454 GS
Explanation - The MinION is a USB‑powered, pocket‑size device that allows sequencing in remote locations with minimal infrastructure.
Correct answer is: Oxford Nanopore MinION
Q.26 Why is 'GC bias' a concern in Illumina sequencing libraries?
GC‑rich regions are difficult to fragment
GC‑rich regions amplify poorly during PCR
GC‑rich regions produce longer reads
GC‑rich regions cannot be sequenced on a flow cell
Explanation - During PCR amplification, high GC content can cause reduced amplification efficiency, resulting in underrepresentation of those regions.
Correct answer is: GC‑rich regions amplify poorly during PCR
Q.27 Which of the following best describes a 'de‑novo' assembly?
Assembling reads against a reference genome
Assembling reads to reconstruct a genome from scratch
Assembling only coding sequences
Assembling transcripts from RNA‑seq data
Explanation - De‑novo assembly constructs a genome using only the sequence reads, without relying on an existing reference.
Correct answer is: Assembling reads to reconstruct a genome from scratch
Q.28 In what way does 'error correction' improve long‑read assemblies?
It shortens read lengths to match Illumina reads
It removes low‑quality reads from the dataset
It uses high‑accuracy short reads to polish long‑read sequences
It replaces homopolymer runs with consensus sequences
Explanation - Hybrid error correction combines the high accuracy of short reads (e.g., Illumina) with long reads to correct errors and produce more accurate assemblies.
Correct answer is: It uses high‑accuracy short reads to polish long‑read sequences
Q.29 What is a key benefit of using a 'dual‑indexing' strategy in Illumina libraries?
Increases read length
Reduces sequencing cost by reusing flow cells
Allows multiplexing of many samples with unique index combinations
Improves base‑calling accuracy
Explanation - Dual‑indexing adds unique barcodes on both ends of each fragment, enabling many samples to be sequenced simultaneously while preventing mis‑assignment.
Correct answer is: Allows multiplexing of many samples with unique index combinations
Q.30 Which of the following best describes the 'phred‑score' in Illumina FASTQ files?
The probability that a base call is correct
The quality of the fluorescent signal
The probability that a base call is incorrect
The number of cycles in a sequencing run
Explanation - Phred‑score is calculated as Q = –10 log10(p), where p is the probability of a wrong base call.
Correct answer is: The probability that a base call is incorrect
Q.31 Which technology is known for 'real‑time' data generation?
Illumina NovaSeq
Ion Torrent Proton
PacBio Sequel II
Oxford Nanopore MinION
Explanation - Nanopore sequencing streams data as nucleotides pass through the pore, allowing immediate analysis.
Correct answer is: Oxford Nanopore MinION
Q.32 In Illumina sequencing, the term 'cluster' refers to:
A group of sequencing reads from the same sample
A group of identical DNA fragments amplified on the flow‑cell surface
A region of high GC content
A set of paired‑end reads
Explanation - Clusters are formed by bridge amplification and contain many copies of the same DNA fragment, enabling high‑sensitivity base calling.
Correct answer is: A group of identical DNA fragments amplified on the flow‑cell surface
Q.33 Why is 'polymerase read length' important in PacBio sequencing?
It determines how many times the polymerase can cycle around a circular template
It limits the number of reads in a run
It sets the maximum read length that can be obtained
It dictates the base‑calling speed
Explanation - The polymerase read length defines how long a continuous stretch can be sequenced before the polymerase dissociates, affecting overall read length.
Correct answer is: It determines how many times the polymerase can cycle around a circular template
Q.34 Which of the following is NOT a common cause of base‑calling errors in Illumina sequencing?
Phasing and pre‑phasing errors
Over‑clustering of reads
DNA damage during library prep
Inaccurate base‑calling algorithms
Explanation - While DNA damage can affect sequencing, Illumina errors are typically due to phasing, clustering, or software miscalls.
Correct answer is: DNA damage during library prep
Q.35 The main function of a 'barcode' in sequencing library preparation is to:
Increase read length
Reduce sequencing errors
Allow samples to be multiplexed in a single run
Enable real‑time analysis
Explanation - Barcodes uniquely label individual samples so they can be pooled and later separated computationally.
Correct answer is: Allow samples to be multiplexed in a single run
Q.36 What does the abbreviation 'QC' stand for in the context of sequencing data analysis?
Quality Control
Quick Check
Sequencing Quality
Read Quality
Explanation - QC refers to procedures used to assess and improve the quality of raw sequencing data before downstream analysis.
Correct answer is: Quality Control
Q.37 Which sequencing technology uses 'T7 RNA polymerase' to generate RNA for sequencing?
PacBio SMRT
Illumina
Ion Torrent
Roche 454
Explanation - Roche 454 used pyrosequencing, which relied on the enzyme T7 RNA polymerase for signal generation.
Correct answer is: Roche 454
Q.38 In the context of NGS, 'adapter trimming' is performed to:
Increase the read length
Remove non‑genomic sequences that can hinder alignment
Add barcodes to each read
Improve base‑calling accuracy
Explanation - Trimming removes adapter and low‑quality tail sequences so that alignment algorithms can map reads more accurately.
Correct answer is: Remove non‑genomic sequences that can hinder alignment
Q.39 Which of the following platforms historically used pyrosequencing?
Illumina HiSeq
Roche 454 GS
PacBio RS
Ion Torrent PGM
Explanation - Roche 454 GS was the first platform to use pyrosequencing, detecting light emitted when pyrophosphate is released during nucleotide addition.
Correct answer is: Roche 454 GS
Q.40 Why do long‑read technologies often require more input DNA than short‑read methods?
Long reads require more sequencing cycles
Long reads are generated by more expensive reagents
Long‑read sequencing reactions are less efficient and need higher template amounts to achieve sufficient coverage
Short reads require less input because they are more accurate
Explanation - Long‑read platforms typically have lower throughput per unit DNA, so more input is needed to reach a comparable depth.
Correct answer is: Long‑read sequencing reactions are less efficient and need higher template amounts to achieve sufficient coverage
Q.41 Which of the following best explains 'phasing' in the context of Illumina sequencing chemistry?
The alignment of reads to a reference genome
The synchronization of nucleotide incorporation across DNA strands in a cluster
The loss of fluorescence signal over time
The process of sequencing both strands of a duplex DNA
Explanation - Phasing occurs when different strands in a cluster incorporate nucleotides at slightly different rates, leading to a mixture of signals.
Correct answer is: The synchronization of nucleotide incorporation across DNA strands in a cluster
Q.42 In what scenario is a 'de‑novo assembly' most likely to be preferred?
Sequencing a well‑annotated reference genome
Assembling a viral genome from clinical samples
Mapping RNA‑seq reads to a reference transcriptome
Calling SNPs in a human exome
Explanation - If no reference is available or the virus has many divergent strains, de‑novo assembly provides an unbiased reconstruction.
Correct answer is: Assembling a viral genome from clinical samples
Q.43 Which of the following metrics is NOT typically used to assess sequencing data quality?
Mean Phred score
GC content distribution
Read duplication rate
Sequencing temperature
Explanation - Temperature is a parameter of the instrument, not a metric for evaluating data quality.
Correct answer is: Sequencing temperature
Q.44 Which of these sequencing platforms is known for a 'zero‑error' consensus read mode?
PacBio Sequel II
Oxford Nanopore PromethION
Illumina NovaSeq
Ion Torrent PGM
Explanation - PacBio’s circular consensus sequencing (CCS) can produce sub‑1% error rates, effectively creating error‑free reads.
Correct answer is: PacBio Sequel II
Q.45 What does the term 'throughput' most directly influence in a sequencing experiment?
Read length
Base‑calling accuracy
Sequencing speed and cost per sample
Library preparation complexity
Explanation - Higher throughput means more data per run, allowing multiple samples to be processed faster and cheaper.
Correct answer is: Sequencing speed and cost per sample
Q.46 Which of the following best describes a 'paired‑end read'?
Two reads that share a common barcode
Two reads that come from opposite ends of the same DNA fragment
A read that contains two different sequencing tags
Two reads that are sequenced in the same cycle
Explanation - Paired‑end sequencing captures information from both ends of a fragment, aiding in structural variant detection.
Correct answer is: Two reads that come from opposite ends of the same DNA fragment
Q.47 Which of the following is a typical advantage of 'real‑time' sequencing?
Improved read accuracy
Instant data availability for rapid decision making
Higher throughput
Lower sequencing cost
Explanation - Real‑time sequencing generates data as it runs, enabling immediate analysis and quicker responses, especially in clinical or field settings.
Correct answer is: Instant data availability for rapid decision making
Q.48 What is the primary function of the 'DNA polymerase' in Illumina sequencing?
To generate fluorescent signals
To incorporate reversible terminators during sequencing by synthesis
To ligate adapters to DNA fragments
To separate DNA strands
Explanation - DNA polymerase adds nucleotides to the growing strand while the terminator prevents further addition until the signal is read.
Correct answer is: To incorporate reversible terminators during sequencing by synthesis
Q.49 Which of the following is a challenge associated with high GC content regions in sequencing?
They are over‑represented in the final data
They cause the polymerase to stall, leading to low coverage
They produce too many adapter sequences
They increase read length beyond the platform limits
Explanation - High GC content can form stable secondary structures that impede polymerase progression, reducing coverage in those regions.
Correct answer is: They cause the polymerase to stall, leading to low coverage
Q.50 The term 'multiplexing' in sequencing refers to:
Sequencing multiple samples in a single run using unique barcodes
Sequencing both strands of DNA simultaneously
Sequencing very long reads
Sequencing without adapters
Explanation - Multiplexing combines several indexed libraries, reducing costs and increasing sample throughput.
Correct answer is: Sequencing multiple samples in a single run using unique barcodes
Q.51 Which sequencing platform is primarily used for ultra‑long read applications such as telomere sequencing?
Illumina NovaSeq
PacBio Sequel II
Oxford Nanopore MinION
Ion Torrent S5
Explanation - Nanopore can generate reads exceeding 50 kb, making it ideal for resolving telomeric repeats.
Correct answer is: Oxford Nanopore MinION
Q.52 What does the 'k‑mer' analysis of a sequencing dataset help to determine?
The size of the flow cell
The distribution of GC content
The estimated genome size and heterozygosity
The error rate of the sequencing platform
Explanation - K‑mer frequency histograms provide estimates of genome size, repeat content, and heterozygosity before assembly.
Correct answer is: The estimated genome size and heterozygosity
Q.53 Which of the following best describes 'coverage uniformity'?
How evenly reads are distributed across the genome
The maximum read length achieved
The total number of reads generated
The base‑calling accuracy across different regions
Explanation - Coverage uniformity assesses whether sequencing depth is consistent across all genomic regions, which is crucial for accurate variant calling.
Correct answer is: How evenly reads are distributed across the genome
Q.54 In Illumina sequencing, what does the term 'cycle' refer to?
A complete sequencing run
The addition of one type of fluorescently labeled nucleotide
The read of one base in a single read
The time it takes to finish a run
Explanation - Each cycle introduces one of four nucleotides; after 150 cycles, reads can be 150 bp long.
Correct answer is: The addition of one type of fluorescently labeled nucleotide
Q.55 What is a major advantage of using 'duplex sequencing' in NGS?
Increases read length
Reduces sequencing errors by confirming both DNA strands
Allows for higher throughput
Simplifies library preparation
Explanation - Duplex sequencing tags each strand uniquely and requires confirmation of the same mutation on both strands to call a variant, dramatically lowering error rates.
Correct answer is: Reduces sequencing errors by confirming both DNA strands
Q.56 Which of the following best describes an 'index hopping' artifact?
When the sequencing index is lost during base calling
When reads are incorrectly assigned to the wrong sample due to index mis‑assignment
When the flow cell temperature changes
When the sequencing run is interrupted
Explanation - Index hopping refers to the erroneous transfer of barcodes between DNA fragments, leading to sample contamination.
Correct answer is: When reads are incorrectly assigned to the wrong sample due to index mis‑assignment
Q.57 In a sequencing run, the term 'base calling' specifically refers to:
Adding adapters to the DNA fragments
Predicting which nucleotide is present at each position in a read
Sorting reads by quality score
Mapping reads to a reference genome
Explanation - Base calling is the computational step that assigns A, C, G, or T to each fluorescence signal detected.
Correct answer is: Predicting which nucleotide is present at each position in a read
Q.58 Which of the following is a common application for Ion Torrent sequencing?
Large‑scale human genome sequencing
Targeted sequencing of cancer gene panels
Full‑length transcript sequencing
Environmental microbiome profiling
Explanation - Ion Torrent's rapid, cost‑effective runs are ideal for targeted panels in clinical diagnostics.
Correct answer is: Targeted sequencing of cancer gene panels
Q.59 Which metric is most informative for evaluating 'read quality' in an Illumina FASTQ file?
Mean read length
Mean Phred quality score
GC content
Number of adapter sequences
Explanation - The Phred score directly indicates the probability of an incorrect base call, summarizing overall read quality.
Correct answer is: Mean Phred quality score
Q.60 Why does the 'error rate' tend to increase toward the end of Illumina reads?
The polymerase becomes less processive
Fluorescent signal fades and becomes harder to distinguish
The DNA template degrades over time
The flow cell surface deteriorates
Explanation - Signal intensity decreases in later cycles, causing more base‑calling errors.
Correct answer is: Fluorescent signal fades and becomes harder to distinguish
Q.61 Which of the following best describes the 'FASTQ' file format?
A compressed binary format for raw sequencing data
A text format that stores nucleotide sequences and associated quality scores
A tabular format for variant calls
A proprietary format used by Illumina only
Explanation - FASTQ files contain the read sequence and a corresponding Phred quality score for each base.
Correct answer is: A text format that stores nucleotide sequences and associated quality scores
Q.62 In the context of NGS, what is 'adapter ligation'?
Attaching a polymerase enzyme to DNA fragments
Adding short DNA sequences to the ends of DNA fragments for amplification and sequencing
Connecting two DNA strands together
Inserting barcodes into the center of a read
Explanation - Adapters provide priming sites for amplification, indexing, and flow‑cell attachment.
Correct answer is: Adding short DNA sequences to the ends of DNA fragments for amplification and sequencing
Q.63 Which of the following technologies uses 'solid‑state' nanopores to detect DNA sequences?
PacBio Sequel II
Ion Torrent PGM
Oxford Nanopore MinION
Illumina NovaSeq
Explanation - Nanopore sequencing measures changes in ionic current as DNA passes through a solid‑state or protein pore.
Correct answer is: Oxford Nanopore MinION
Q.64 The 'Read 1' and 'Read 2' designations in Illumina sequencing refer to:
The first and second cycles of base calling
The two strands of the same DNA molecule
Two separate sequencing lanes on the flow cell
The two paired‑end reads from each DNA fragment
Explanation - Read 1 and Read 2 capture sequences from opposite ends of the same fragment in a paired‑end run.
Correct answer is: The two paired‑end reads from each DNA fragment
Q.65 What is the purpose of 'size selection' in a sequencing library?
To remove short adapter fragments and ensure uniform read length distribution
To increase the number of PCR duplicates
To improve base‑calling accuracy by selecting longer reads
To select only high‑GC regions for sequencing
Explanation - Size selection eliminates unwanted fragments like adapters and very short pieces, improving library complexity.
Correct answer is: To remove short adapter fragments and ensure uniform read length distribution
Q.66 Which sequencing platform uses 'time‑of‑flight' mass spectrometry for base detection?
Illumina
PacBio
Ion Torrent
Roche 454
Explanation - Ion Torrent detects the release of hydrogen ions during nucleotide incorporation, which is a form of mass change.
Correct answer is: Ion Torrent
Q.67 Which of the following best describes 'homopolymer run errors'?
Missing bases at the start of a read
Incorrectly counting the number of bases in a stretch of identical nucleotides
Phasing errors in Illumina sequencing
Errors due to GC content bias
Explanation - Platforms like Ion Torrent and early Illumina versions struggle to determine the exact length of homopolymer tracts.
Correct answer is: Incorrectly counting the number of bases in a stretch of identical nucleotides
Q.68 What does the term 'sequencing depth' mean in genomics?
The number of nucleotides sequenced in a single read
The average number of times each base in the genome is read
The maximum number of samples sequenced in one run
The level of coverage across the entire genome
Explanation - Sequencing depth, or coverage, is defined as how many times a given base is sequenced on average.
Correct answer is: The average number of times each base in the genome is read
Q.69 In Illumina sequencing, why is the term 'flow cell' used?
It indicates the path where the DNA flows through the machine
It is the chamber where the sequencing reactions occur, containing flow‑cell channels
It is the name of the software controlling the instrument
It refers to the step where DNA is flowed onto the sequencing surface
Explanation - A flow cell is a chip with microchannels where clusters are amplified and sequenced.
Correct answer is: It is the chamber where the sequencing reactions occur, containing flow‑cell channels
Q.70 Which of the following sequencing technologies is best suited for detecting structural variants in a genome?
Illumina short reads
PacBio long reads
Ion Torrent short reads
Roche 454 pyrosequencing
Explanation - Long reads can span large repeats and complex regions, enabling more accurate structural variant detection.
Correct answer is: PacBio long reads
Q.71 What does the abbreviation 'PCR' stand for?
Polymerase Chain Reaction
Protein Coding Region
Pooled Clone Repository
Phased Chromatin Repertoire
Explanation - PCR is a laboratory technique used to amplify DNA segments, commonly used in library preparation.
Correct answer is: Polymerase Chain Reaction
Q.72 Why is a 'reference genome' important in NGS data analysis?
It allows mapping of reads to known coordinates for variant detection
It provides a universal primer for PCR amplification
It eliminates the need for quality control
It ensures all reads are of high quality
Explanation - A reference genome provides the baseline sequence against which reads are aligned to identify differences.
Correct answer is: It allows mapping of reads to known coordinates for variant detection
Q.73 Which of the following is a major benefit of using 'duplex consensus sequencing'?
Higher throughput
Lower cost per base
Improved error correction by requiring concordant signals on both strands
Shorter read lengths
Explanation - Duplex consensus requires that a variant be seen on both strands to be called, greatly reducing false positives.
Correct answer is: Improved error correction by requiring concordant signals on both strands
Q.74 In Nanopore sequencing, the 'mean current drop' is used to infer:
The length of the read
The type of nucleotide being translocated
The speed of the motor protein
The temperature of the sequencing reaction
Explanation - Different nucleotides produce distinct current signatures as they pass through the pore.
Correct answer is: The type of nucleotide being translocated
Q.75 Which sequencing platform is known for having the highest per‑run throughput in 2025?
Illumina NovaSeq 6000
Oxford Nanopore PromethION 48‑channel
PacBio Sequel IIe
Ion Torrent S5
Explanation - The NovaSeq 6000 can produce up to 6 TB of data per run, making it the highest‑throughput system available.
Correct answer is: Illumina NovaSeq 6000
Q.76 What is the main drawback of using a 'PCR‑based' library preparation method?
It generates longer reads
It introduces amplification bias and can create duplicate reads
It eliminates adapter contamination
It reduces sequencing cost
Explanation - PCR amplification can preferentially amplify some fragments, leading to uneven representation and duplicate reads.
Correct answer is: It introduces amplification bias and can create duplicate reads
Q.77 Which sequencing technology uses 'flow‑cell tiles' for spatial separation of clusters?
Illumina HiSeq
PacBio RS II
Oxford Nanopore MinION
Roche 454 GS
Explanation - Illumina flow cells are partitioned into tiles to allow high‑density cluster imaging and sequencing.
Correct answer is: Illumina HiSeq
Q.78 In the context of next‑generation sequencing, what does 'single‑cell sequencing' refer to?
Sequencing the genome of an entire organism in one run
Sequencing the DNA of an individual cell, often to study cellular heterogeneity
Sequencing only one type of DNA fragment per run
Sequencing the genome of a single individual
Explanation - Single‑cell sequencing allows researchers to examine genetic variations among individual cells in a population.
Correct answer is: Sequencing the DNA of an individual cell, often to study cellular heterogeneity
Q.79 Which of the following best explains why Illumina sequencing requires a 'phased' sequencing cycle?
To ensure all polymerases are synchronized during base incorporation
To separate the flow cell into multiple tiles
To increase read length
To prevent chemical cross‑linking
Explanation - Phasing ensures that all DNA strands in a cluster incorporate nucleotides simultaneously for accurate signal detection.
Correct answer is: To ensure all polymerases are synchronized during base incorporation
Q.80 The 'library complexity' of a sequencing prep refers to:
The diversity of unique DNA fragments in the library
The average read length
The number of PCR cycles performed
The GC content distribution
Explanation - High library complexity means many unique fragments, which reduces duplication and improves coverage.
Correct answer is: The diversity of unique DNA fragments in the library
Q.81 Which sequencing method relies on the detection of fluorescent signals emitted by a laser?
Illumina sequencing
PacBio sequencing
Oxford Nanopore sequencing
Ion Torrent sequencing
Explanation - Illumina uses a laser to excite fluorescent tags on nucleotides for base detection.
Correct answer is: Illumina sequencing
Q.82 Why is 'base calling' more challenging for Nanopore sequencing compared to Illumina?
Nanopore produces longer reads
Nanopore signals are noisy and require complex algorithms for base calling
Illumina does not require base calling
Nanopore uses a different type of polymerase
Explanation - The ionic current signal is less discrete than fluorescent signals, requiring sophisticated computational models.
Correct answer is: Nanopore signals are noisy and require complex algorithms for base calling
Q.83 What is the primary purpose of a 'barcode' in a multiplexed sequencing run?
To increase read length
To uniquely identify each sample within a pool
To reduce sequencing errors
To attach adapters to DNA fragments
Explanation - Barcodes allow multiple libraries to be sequenced together and later computationally separated.
Correct answer is: To uniquely identify each sample within a pool
Q.84 Which sequencing platform uses a 'solid‑state' nanopore technology?
Oxford Nanopore MinION
Illumina NovaSeq
PacBio Sequel II
Ion Torrent S5
Explanation - Nanopore sequencing measures electrical current changes as DNA passes through a nanopore.
Correct answer is: Oxford Nanopore MinION
Q.85 In Illumina sequencing, what is the typical number of cycles per read in a 150‑bp run?
50
100
150
200
Explanation - Each cycle adds one base, so a 150‑bp read requires 150 cycles.
Correct answer is: 150
Q.86 Which of the following best describes the 'error profile' of a sequencing technology?
The number of reads per run
The distribution of sequencing errors across read length and sequence context
The cost per megabase
The maximum read length
Explanation - Error profile includes types of errors (substitutions, indels) and where they are most likely to occur.
Correct answer is: The distribution of sequencing errors across read length and sequence context
Q.87 What does 'k‑mer' refer to in genomics?
A cluster of DNA fragments in a flow cell
A sequence of k consecutive nucleotides
A type of sequencing error
A length‑based quality metric
Explanation - K‑mers are substrings of length k extracted from sequences, useful for assembly and error correction.
Correct answer is: A sequence of k consecutive nucleotides
Q.88 Which of the following technologies is best suited for real‑time, portable sequencing in the field?
Illumina NovaSeq
PacBio Sequel II
Oxford Nanopore MinION
Roche 454
Explanation - The MinION is a small, USB‑powered device that can sequence DNA in real time, ideal for field use.
Correct answer is: Oxford Nanopore MinION
Q.89 Which sequencing platform uses 'pulsed electrochemical detection' to measure nucleotide incorporation?
Ion Torrent
Illumina
PacBio
Oxford Nanopore
Explanation - Ion Torrent detects hydrogen ions released during nucleotide addition, creating an electrochemical pulse.
Correct answer is: Ion Torrent
Q.90 What does the 'Q30' quality threshold indicate in Illumina sequencing?
30% of reads are longer than 30 bp
A 1 in 1000 chance of an incorrect base call
A 30 % probability of a sequencing error
Reads are 30 bp long
Explanation - Q30 corresponds to a base‑calling error probability of 0.1% (1/1000).
Correct answer is: A 1 in 1000 chance of an incorrect base call
Q.91 Which of the following is a typical use of 'mate‑pair sequencing'?
Detecting short SNPs
Resolving large structural variants and scaffolding genomes
Performing RNA‑seq
Targeted gene panel sequencing
Explanation - Mate‑pair libraries capture long insert sizes (2–5 kb), aiding in assembly and variant detection.
Correct answer is: Resolving large structural variants and scaffolding genomes
Q.92 Which sequencing technology uses 'time‑of‑flight mass spectrometry' for nucleotide detection?
Illumina
PacBio
Ion Torrent
Roche 454
Explanation - Ion Torrent measures the release of hydrogen ions, which is a mass change detected by the system.
Correct answer is: Ion Torrent
Q.93 In Nanopore sequencing, what is a 'translocation' event?
The movement of a DNA polymerase across the template
The movement of a DNA fragment through the nanopore
The conversion of a DNA strand into RNA
The ligation of adapters to DNA fragments
Explanation - Translocation refers to the passage of DNA through the pore, generating the electrical signal.
Correct answer is: The movement of a DNA fragment through the nanopore
Q.94 Which of the following is a key benefit of 'PCR‑free' library preparation?
Higher error rates
Reduced bias and improved representation of GC‑rich regions
Simplified workflow
Lower input DNA requirement
Explanation - PCR amplification can skew representation; PCR‑free preserves the original sequence composition.
Correct answer is: Reduced bias and improved representation of GC‑rich regions
Q.95 Which of the following technologies uses 'laser‑based imaging' to detect base incorporation?
Illumina sequencing
PacBio sequencing
Oxford Nanopore sequencing
Ion Torrent sequencing
Explanation - Illumina uses a laser to excite fluorescent tags during sequencing‑by‑synthesis.
Correct answer is: Illumina sequencing
Q.96 Which sequencing technology was the first to use 'single‑molecule real‑time' (SMRT) sequencing?
PacBio Sequel
Ion Torrent PGM
Illumina NextSeq
Oxford Nanopore MinION
Explanation - PacBio’s SMRT technology records DNA synthesis in real time at the single‑molecule level.
Correct answer is: PacBio Sequel
Q.97 What is the main source of error in Illumina sequencing known as 'phasing'?
Inaccurate base calling due to low signal intensity
Misalignment of reads to the reference genome
Strand synthesis not synchronized across clusters
Loss of cluster information during imaging
Explanation - Phasing occurs when DNA strands in a cluster incorporate nucleotides at different times, leading to mixed signals.
Correct answer is: Strand synthesis not synchronized across clusters
Q.98 Which sequencing platform typically uses a 'tethered DNA polymerase' to generate reads?
PacBio Sequel
Illumina HiSeq
Ion Torrent Proton
Oxford Nanopore PromethION
Explanation - PacBio attaches the polymerase to the surface via a tether, enabling long read acquisition.
Correct answer is: PacBio Sequel
Q.99 In the context of NGS, what does 'adapter contamination' most commonly lead to?
Improved read quality
Incorrect read mapping and variant calling errors
Shorter reads
Increased coverage uniformity
Explanation - Untrimmed adapters can interfere with alignment, causing mis‑alignments and false variants.
Correct answer is: Incorrect read mapping and variant calling errors
Q.100 Which of the following best explains why Ion Torrent sequencing struggles with homopolymers?
It relies on fluorescent tags that cannot distinguish repeats
It measures the number of hydrogen ions released, which can saturate the signal for multiple identical nucleotides
It cannot read GC‑rich sequences
It requires PCR amplification of homopolymer regions
Explanation - The pH signal can’t differentiate between 4 and 5 of the same base, leading to indel errors.
Correct answer is: It measures the number of hydrogen ions released, which can saturate the signal for multiple identical nucleotides
Q.101 Which of the following sequencing strategies is used for capturing all exonic regions of the genome?
Whole genome sequencing
Targeted exome sequencing
RNA‑seq
Single‑cell sequencing
Explanation - Exome capture targets exons for sequencing, enabling efficient variant detection in coding regions.
Correct answer is: Targeted exome sequencing
Q.102 Which of the following best describes a 'dual index' in Illumina library prep?
Two unique barcodes added to each fragment, one on each end
A pair of indices added to the sequencing flow cell
Two sets of adapter sequences
Two reference genomes used for alignment
Explanation - Dual indexing reduces mis‑assignment and allows for higher sample multiplexing.
Correct answer is: Two unique barcodes added to each fragment, one on each end
Q.103 Which of the following is NOT an advantage of long‑read sequencing?
Improved detection of structural variants
Simplified genome assembly
Higher accuracy per base
Ability to span repetitive regions
Explanation - Long‑read platforms typically have higher raw error rates than short‑read platforms.
Correct answer is: Higher accuracy per base
Q.104 What is the primary function of a 'flow cell' in Illumina sequencing?
To amplify DNA fragments via PCR
To provide a surface where DNA clusters can form and be imaged
To store reagents during sequencing
To separate different DNA libraries
Explanation - The flow cell contains millions of nanowells where clusters amplify and are imaged during sequencing.
Correct answer is: To provide a surface where DNA clusters can form and be imaged
Q.105 Which sequencing platform is known for producing 'single‑end' reads only?
PacBio Sequel II
Illumina MiSeq
Ion Torrent PGM
Oxford Nanopore MinION
Explanation - The PGM and many older Ion Torrent models generate only single‑end reads, though newer ones support paired‑end.
Correct answer is: Ion Torrent PGM
Q.106 What does 'adapter ligation' accomplish in a sequencing library?
It adds primers for PCR amplification
It attaches synthetic DNA sequences to enable sequencing and indexing
It fragments the DNA sample
It repairs damaged DNA ends
Explanation - Adapters provide priming sites for sequencing chemistry and often contain barcodes.
Correct answer is: It attaches synthetic DNA sequences to enable sequencing and indexing
Q.107 Which of the following best explains why Nanopore sequencing can produce reads exceeding 100 kb?
It uses a very long polymerase enzyme
It employs a large pore that allows many nucleotides to pass through simultaneously
It reads DNA without breaking it into fragments, allowing continuous long reads
It uses a special DNA polymerase that can read through repetitive sequences
Explanation - Nanopore sequencing processes intact DNA molecules, producing very long continuous reads.
Correct answer is: It reads DNA without breaking it into fragments, allowing continuous long reads
Q.108 Which of the following is a major source of bias in Illumina library preparation?
Fragmentation of DNA
Amplification during PCR
Size selection
Adapter ligation
Explanation - PCR can preferentially amplify fragments based on GC content or length, creating bias.
Correct answer is: Amplification during PCR
Q.109 Which sequencing platform uses 'photons' to detect base incorporation?
Illumina sequencing
PacBio Sequel
Oxford Nanopore MinION
Ion Torrent PGM
Explanation - Illumina’s sequencing‑by‑synthesis uses fluorescently labeled nucleotides that emit photons when incorporated.
Correct answer is: Illumina sequencing
Q.110 In the context of DNA sequencing, what does the term 'read length' refer to?
The total number of reads generated
The average number of times a base is read
The number of nucleotides in a single sequence read
The duration of a sequencing run
Explanation - Read length is the length, in base pairs, of each sequenced read.
Correct answer is: The number of nucleotides in a single sequence read
Q.111 Which of the following best describes 'coverage uniformity' in sequencing experiments?
The average read length across all samples
The consistency of sequencing depth across different genomic regions
The uniform distribution of sequencing errors
The evenness of GC content across reads
Explanation - Coverage uniformity ensures that all areas of the genome are covered evenly, reducing blind spots.
Correct answer is: The consistency of sequencing depth across different genomic regions
Q.112 Which of the following sequencing methods is most suitable for detecting large structural variants?
Illumina short‑read sequencing
PacBio long‑read sequencing
Ion Torrent short‑read sequencing
Sanger sequencing
Explanation - Long reads can span large repeats and structural variants, making detection more accurate.
Correct answer is: PacBio long‑read sequencing
Q.113 Which of the following is a characteristic of a 'duplex consensus read'?
It is derived from a single DNA strand
It requires a single sequencing read to be accurate
It is formed by combining reads from both strands of a DNA duplex to reduce errors
It is only applicable to Illumina sequencing
Explanation - Duplex sequencing tags each strand uniquely and requires concordant evidence to call a variant, improving accuracy.
Correct answer is: It is formed by combining reads from both strands of a DNA duplex to reduce errors
Q.114 Which sequencing technology uses a 'fluorescence‑based' detection method?
PacBio SMRT
Oxford Nanopore
Illumina sequencing
Ion Torrent
Explanation - Illumina uses fluorescently labeled nucleotides that are detected by a camera during sequencing.
Correct answer is: Illumina sequencing
Q.115 What is the main limitation of 'PCR‑based' library preparation?
High input DNA requirement
Introduction of amplification bias and duplicates
Inability to sequence long reads
Low throughput
Explanation - PCR can preferentially amplify some fragments, leading to uneven coverage and duplicate reads.
Correct answer is: Introduction of amplification bias and duplicates
Q.116 Which of the following sequencing methods is best suited for whole‑genome sequencing of bacteria?
Illumina short‑read sequencing
PacBio long‑read sequencing
Oxford Nanopore MinION
All of the above
Explanation - Each platform can be used for bacterial WGS; the choice depends on budget, desired read length, and turnaround time.
Correct answer is: All of the above
Q.117 What is a common consequence of high GC content in a sequencing library?
Increased read length
Reduced sequencing bias
Lower PCR efficiency and coverage drop‑outs
Improved accuracy
Explanation - GC‑rich regions can form stable structures, hindering amplification and sequencing, leading to lower coverage.
Correct answer is: Lower PCR efficiency and coverage drop‑outs
Q.118 Which of the following best defines 'single‑end sequencing'?
Sequencing both ends of a DNA fragment
Sequencing one end of a DNA fragment only
Sequencing the entire genome in a single run
Sequencing only the exome of a genome
Explanation - Single‑end sequencing generates a single read from one end of each fragment, as opposed to paired‑end.
Correct answer is: Sequencing one end of a DNA fragment only
Q.119 Which of the following best describes a 'barcode' in sequencing?
A short sequence that identifies a sample in multiplexed runs
A type of adapter used for library preparation
A read length measurement
A quality score
Explanation - Barcodes (indices) are unique short sequences that allow pooling of multiple libraries in one sequencing run.
Correct answer is: A short sequence that identifies a sample in multiplexed runs
Q.120 Which of the following sequencing platforms relies on the detection of changes in ionic current?
Illumina HiSeq
PacBio Sequel
Oxford Nanopore MinION
Ion Torrent PGM
Explanation - Nanopore sequencing measures ionic current changes as DNA passes through a nanopore.
Correct answer is: Oxford Nanopore MinION
Q.121 What does the term 'read duplication rate' indicate?
The proportion of reads that are identical due to PCR duplicates or high library complexity
The speed of the sequencing instrument
The number of reads that failed quality control
The proportion of reads with adapter contamination
Explanation - High duplication indicates PCR bias or low library diversity, reducing effective coverage.
Correct answer is: The proportion of reads that are identical due to PCR duplicates or high library complexity
Q.122 Which of the following is a characteristic of Ion Torrent sequencing?
Uses fluorescent labels for base detection
Measures hydrogen ion release during nucleotide incorporation
Requires a flow cell with thousands of clusters
Generates long reads over 10 kb
Explanation - Ion Torrent uses a pH sensor to detect the release of H⁺ ions when nucleotides are incorporated.
Correct answer is: Measures hydrogen ion release during nucleotide incorporation
Q.123 Which of the following is a major challenge in sequencing highly repetitive genomic regions?
They cause the sequencer to overheat
They produce long reads
They create difficulty in read alignment and assembly
They increase base‑calling accuracy
Explanation - Repeats can confuse aligners, leading to mis‑alignment and mis‑assembly of the genome.
Correct answer is: They create difficulty in read alignment and assembly
Q.124 Which sequencing technology typically offers the lowest cost per base?
Illumina NovaSeq
PacBio Sequel II
Oxford Nanopore MinION
Sanger sequencing
Explanation - Illumina’s high throughput yields a low cost per base, making it the most economical for large genomes.
Correct answer is: Illumina NovaSeq
Q.125 Which of the following is an advantage of using a 'dual‑index' in Illumina sequencing?
It reduces sequencing errors
It increases read length
It enables higher multiplexing and reduces index hopping
It eliminates the need for PCR amplification
Explanation - Dual indices provide a unique barcode pair, improving sample identification and reducing mis‑assignment.
Correct answer is: It enables higher multiplexing and reduces index hopping
Q.126 Which of the following sequencing platforms is best for detecting epigenetic modifications such as 5‑methylcytosine?
Illumina sequencing
Oxford Nanopore MinION
PacBio Sequel II
Ion Torrent PGM
Explanation - Nanopore can directly detect modified bases in real time due to altered electrical signals.
Correct answer is: Oxford Nanopore MinION
Q.127 In sequencing, what is meant by the term 'library complexity'?
The diversity of unique fragments in the sequencing library
The number of different sequencing platforms used
The length distribution of the reads
The GC content of the library
Explanation - High library complexity reduces duplication and improves coverage uniformity.
Correct answer is: The diversity of unique fragments in the sequencing library
Q.128 Which sequencing technology was the first to use 'solid‑state' nanopores for DNA sequencing?
Oxford Nanopore MinION
PacBio Sequel
Ion Torrent PGM
Illumina HiSeq
Explanation - Nanopore sequencing uses a protein or solid‑state pore to detect nucleotides as they pass through.
Correct answer is: Oxford Nanopore MinION
Q.129 Which of the following best explains the term 'mean quality score' in a sequencing run?
The average number of reads per sample
The average base‑calling error probability across all reads
The average read length
The average GC content of reads
Explanation - Mean quality score is the average Phred score of all bases, indicating overall sequencing accuracy.
Correct answer is: The average base‑calling error probability across all reads
Q.130 Which sequencing platform typically requires a higher input DNA amount than Illumina?
PacBio Sequel II
Illumina NovaSeq
Oxford Nanopore MinION
Ion Torrent Proton
Explanation - PacBio’s long‑read library prep generally needs more input DNA to reach comparable depth.
Correct answer is: PacBio Sequel II
Q.131 Which of the following is a typical output of a sequencing run?
Fastq files containing read sequences and quality scores
A list of PCR primers
A set of physical DNA fragments
A detailed protocol for library preparation
Explanation - FASTQ files are the standard output format storing sequences and per‑base quality information.
Correct answer is: Fastq files containing read sequences and quality scores
Q.132 Which sequencing technology is most suitable for real‑time pathogen detection in a clinical setting?
Illumina MiSeq
PacBio RS II
Oxford Nanopore MinION
Roche 454
Explanation - Nanopore’s portability and real‑time data output allow rapid identification of pathogens in the clinic.
Correct answer is: Oxford Nanopore MinION
Q.133 What does the term 'library prep' refer to in DNA sequencing?
The process of aligning reads to a reference genome
The generation of a DNA library containing fragments ready for sequencing
The analysis of sequencing data to find variants
The design of primers for PCR amplification
Explanation - Library prep involves fragmentation, end‑repair, adapter ligation, and (sometimes) amplification to prepare DNA for sequencing.
Correct answer is: The generation of a DNA library containing fragments ready for sequencing
Q.134 Which of the following is a key disadvantage of Ion Torrent sequencing?
High error rates in homopolymers
Low throughput
Requirement for large input DNA
Inability to perform paired‑end sequencing
Explanation - The pH‑based detection struggles to count identical nucleotides, leading to indel errors.
Correct answer is: High error rates in homopolymers
Q.135 Which of the following best describes a 'mate‑pair library'?
A library with two different adapters on each fragment
A library containing DNA fragments with large insert sizes that are sequenced from both ends
A library that is amplified in a PCR‑free manner
A library designed for transcriptome analysis
Explanation - Mate‑pair libraries capture long-range information, useful for scaffolding and structural variant detection.
Correct answer is: A library containing DNA fragments with large insert sizes that are sequenced from both ends
Q.136 Which of the following is an advantage of 'single‑molecule' sequencing technologies?
They do not require PCR amplification
They produce only short reads
They have lower error rates than other methods
They require large amounts of input DNA
Explanation - Single‑molecule approaches like PacBio and Nanopore sequence individual DNA molecules, avoiding amplification bias.
Correct answer is: They do not require PCR amplification
Q.137 Which of the following describes the 'flow cell' in Illumina sequencing?
The chip where DNA fragments are amplified into clusters
The software used for base calling
The reagent cartridge that supplies nucleotides
The storage area for sequenced data
Explanation - The flow cell provides a surface with millions of nanowells for cluster formation and imaging.
Correct answer is: The chip where DNA fragments are amplified into clusters
Q.138 Which sequencing technology uses 'time‑of‑flight' detection of hydrogen ions released during DNA synthesis?
Ion Torrent
Illumina
PacBio
Oxford Nanopore
Explanation - Ion Torrent measures the pH change caused by released hydrogen ions after nucleotide incorporation.
Correct answer is: Ion Torrent
Q.139 Which of the following best explains a 'base‑calling' error?
An error in the sequencing machine’s temperature control
Misidentification of the correct nucleotide based on the detection signal
Failure to align reads to the reference genome
Loss of adapter sequences
Explanation - Base‑calling errors arise when the software incorrectly interprets the signal as a different nucleotide.
Correct answer is: Misidentification of the correct nucleotide based on the detection signal
Q.140 What does a 'Q20' score indicate in a sequencing run?
The probability of a base call being incorrect is 1 in 20
The probability of a base call being incorrect is 1 in 200
The probability of a base call being incorrect is 1 in 1000
The probability of a base call being incorrect is 1 in 50
Explanation - A Q20 score corresponds to a 1% error probability (1/100).
Correct answer is: The probability of a base call being incorrect is 1 in 20
Q.141 Which of the following is NOT an advantage of long‑read sequencing platforms?
Ability to resolve repetitive regions
Improved assembly of complex genomes
Higher per‑base accuracy than short reads
Detection of structural variants
Explanation - Long‑read platforms typically have higher raw error rates compared to short‑read systems.
Correct answer is: Higher per‑base accuracy than short reads
Q.142 Which of the following sequencing platforms uses a 'laser‑based imaging' system to detect base incorporation?
Illumina NextSeq
PacBio Sequel
Oxford Nanopore MinION
Ion Torrent S5
Explanation - Illumina NextSeq employs a laser to excite fluorescently labeled nucleotides during sequencing‑by‑synthesis.
Correct answer is: Illumina NextSeq
Q.143 Which sequencing platform is most suited for ultra‑long reads exceeding 1 megabase?
PacBio Sequel IIe
Oxford Nanopore PromethION 48‑channel
Illumina NovaSeq
Ion Torrent Proton
Explanation - PromethION can produce extremely long reads, sometimes exceeding 1 Mb, ideal for telomere sequencing.
Correct answer is: Oxford Nanopore PromethION 48‑channel
Q.144 Which sequencing platform uses 'reverse‑transcription' to sequence RNA directly?
Illumina NovaSeq
PacBio Sequel II
Oxford Nanopore Direct RNA
Ion Torrent PGM
Explanation - Oxford Nanopore’s Direct RNA sequencing captures native RNA molecules without reverse transcription.
Correct answer is: Oxford Nanopore Direct RNA
Q.145 What is the main disadvantage of 'PCR amplification' in library prep?
It reduces library complexity
It increases read length
It reduces sequencing cost
It eliminates the need for adapters
Explanation - PCR amplification can favor some fragments over others, reducing the number of unique sequences.
Correct answer is: It reduces library complexity
Q.146 Which of the following best describes the 'read length' distribution in Illumina sequencing?
Uniform across all reads
Variable but typically 50–150 bp
Always 200 bp
Always 1000 bp
Explanation - Illumina read lengths are usually set between 50 and 150 bp depending on the run configuration.
Correct answer is: Variable but typically 50–150 bp
Q.147 Which sequencing platform is known for having the lowest per‑base error rate after error correction?
Illumina NovaSeq
PacBio Sequel II
Oxford Nanopore MinION
Ion Torrent S5
Explanation - Illumina's short‑read platforms have the lowest raw error rate (~0.001) compared to other technologies.
Correct answer is: Illumina NovaSeq
Q.148 Which sequencing technology uses 'solid‑state nanopores' and can directly detect modified nucleotides?
Oxford Nanopore MinION
PacBio Sequel II
Illumina MiSeq
Ion Torrent PGM
Explanation - Nanopore sequencing can sense chemical modifications as changes in ionic current.
Correct answer is: Oxford Nanopore MinION
Q.149 What does the term 'library complexity' influence in sequencing?
Sequencing run duration
Number of reads per cycle
Depth of coverage and duplication rate
Signal strength during imaging
Explanation - Higher library complexity reduces duplicate reads and improves effective coverage across the genome.
Correct answer is: Depth of coverage and duplication rate
Q.150 Which of the following is a major limitation of Illumina sequencing for genome assembly?
Short read length limiting the resolution of repetitive regions
High per‑base cost
Low throughput
Requirement for large input DNA amounts
Explanation - Short reads can’t span large repeats, making de‑novo assembly of complex genomes more difficult.
Correct answer is: Short read length limiting the resolution of repetitive regions
Q.151 Which of the following best describes the 'cluster density' on an Illumina flow cell?
The number of clusters per unit area of the flow cell surface
The number of reads per base in a sequencing run
The length of the read produced
The percentage of reads that contain adapter sequences
Explanation - Cluster density determines how many DNA molecules are amplified and imaged per square millimeter.
Correct answer is: The number of clusters per unit area of the flow cell surface
Q.152 In Illumina sequencing, which step directly generates fluorescent signal for base detection?
Adapter ligation
Bridge amplification
Sequencing by synthesis with reversible terminators
PCR amplification
Explanation - During each cycle, a fluorescently labeled nucleotide is incorporated, emitting a detectable signal.
Correct answer is: Sequencing by synthesis with reversible terminators
Q.153 Which sequencing platform is most suitable for rapid, field‑based sequencing of environmental samples?
Illumina MiSeq
PacBio Sequel II
Oxford Nanopore MinION
Ion Torrent Proton
Explanation - The MinION’s portability and real‑time output make it ideal for on‑site sequencing.
Correct answer is: Oxford Nanopore MinION
Q.154 What is a major benefit of 'single‑molecule real‑time' (SMRT) sequencing?
It eliminates the need for PCR amplification
It provides extremely short reads
It uses a very low input DNA amount
It requires a flow cell with millions of clusters
Explanation - SMRT sequencing reads individual DNA molecules directly, avoiding PCR bias and duplication.
Correct answer is: It eliminates the need for PCR amplification
Q.155 Which sequencing technology uses a 'solid‑state' pore to generate electrical signals as DNA passes through?
Oxford Nanopore MinION
PacBio Sequel II
Illumina HiSeq
Ion Torrent PGM
Explanation - Nanopore sequencing measures changes in electrical current as DNA passes through a nanopore.
Correct answer is: Oxford Nanopore MinION
Q.156 Which of the following is a key feature of Ion Torrent sequencing?
Uses fluorescently labeled nucleotides
Detects hydrogen ion release via pH changes
Requires a flow cell with clusters
Produces reads longer than 10 kb
Explanation - Ion Torrent measures the pH shift caused by the incorporation of nucleotides.
Correct answer is: Detects hydrogen ion release via pH changes
Q.157 What is the purpose of a 'PCR duplicate filter' during data processing?
To remove reads that are too short
To remove reads containing adapter sequences
To remove identical reads that result from PCR amplification
To correct base‑calling errors
Explanation - Duplicate filtering eliminates over‑represented reads that could bias downstream analysis.
Correct answer is: To remove identical reads that result from PCR amplification
Q.158 Which of the following best explains a 'coverage gap' in sequencing?
A region of the genome not covered by reads, often due to bias or low sequencing depth
A region of high coverage beyond the desired depth
A read that fails quality control
An area where sequencing errors are high
Explanation - Coverage gaps indicate missing data in particular genomic loci, which can hinder variant detection.
Correct answer is: A region of the genome not covered by reads, often due to bias or low sequencing depth
Q.159 Which of the following sequencing platforms uses 'laser‑excited fluorescent detection' during sequencing?
Illumina sequencing
PacBio SMRT
Oxford Nanopore MinION
Ion Torrent PGM
Explanation - Illumina employs a laser to excite the fluorescent dyes attached to nucleotides for base calling.
Correct answer is: Illumina sequencing
Q.160 Which of the following best describes the 'library preparation' step in sequencing?
The process of aligning reads to a reference genome
The process of fragmenting DNA, ligating adapters, and optionally amplifying to create a sequencing library
The step that generates the final FASTQ files
The process of selecting reads based on quality score
Explanation - Library preparation converts raw DNA into a form compatible with sequencing platforms.
Correct answer is: The process of fragmenting DNA, ligating adapters, and optionally amplifying to create a sequencing library
Q.161 In the context of Illumina sequencing, what does 'index hopping' refer to?
The movement of adapters along a DNA strand
The misassignment of read indices between different DNA fragments
The loss of fluorescent signal during base incorporation
The migration of clusters across the flow cell
Explanation - Index hopping occurs when indices (barcodes) are incorrectly transferred, causing sample cross‑contamination.
Correct answer is: The misassignment of read indices between different DNA fragments
Q.162 Which of the following best explains why 'GC bias' can affect sequencing data?
High GC regions produce brighter fluorescence signals
GC‑rich areas are more likely to form secondary structures that inhibit amplification and sequencing
GC content has no effect on sequencing
GC‑rich regions are sequenced more accurately
Explanation - GC‑rich sequences can impede polymerase progress and PCR, leading to uneven coverage.
Correct answer is: GC‑rich areas are more likely to form secondary structures that inhibit amplification and sequencing
Q.163 Which of the following platforms uses 'phospholinked DNA polymerase' to generate long reads?
PacBio Sequel II
Illumina NovaSeq
Oxford Nanopore MinION
Ion Torrent Proton
Explanation - PacBio attaches the polymerase to the DNA via phospholinked anchors, enabling continuous sequencing.
Correct answer is: PacBio Sequel II
Q.164 Which sequencing method is most suitable for sequencing of small genomes (e.g., bacteria) with rapid turnaround?
Illumina MiSeq
PacBio Sequel II
Oxford Nanopore MinION
Ion Torrent PGM
Explanation - Nanopore’s speed and portability make it ideal for quick bacterial genome sequencing.
Correct answer is: Oxford Nanopore MinION
Q.165 In the context of sequencing, what is a 'phasing error'?
An error in aligning reads to a reference genome
An error due to the mis‑synchronization of nucleotides incorporated by polymerase across clusters
An error caused by adapter contamination
An error in the flow cell temperature control
Explanation - Phasing errors occur when some strands lag behind others in the sequencing cycles, leading to mixed signals.
Correct answer is: An error due to the mis‑synchronization of nucleotides incorporated by polymerase across clusters
Q.166 Which sequencing platform is known for having the highest read depth per run?
Illumina NovaSeq
PacBio Sequel II
Oxford Nanopore PromethION
Ion Torrent S5
Explanation - NovaSeq can generate up to 6 TB of data per run, providing the highest depth among platforms.
Correct answer is: Illumina NovaSeq
Q.167 Which of the following is an advantage of 'duplex sequencing' in NGS?
Reduced sequencing cost
Reduced duplication
Lower error rates by validating reads from both strands
Increased read length
Explanation - Duplex sequencing requires both strands to agree, drastically reducing false variant calls.
Correct answer is: Lower error rates by validating reads from both strands
Q.168 Which of the following best describes the 'library complexity' metric?
The number of unique DNA fragments in a sequencing library
The average read length of a sequencing run
The GC content of the library
The number of barcodes used in the library
Explanation - Library complexity refers to how many distinct fragments are present, impacting duplication rates and coverage.
Correct answer is: The number of unique DNA fragments in a sequencing library
Q.169 What is a 'mate‑pair read' commonly used for?
To improve sequencing speed
To span large insert sizes for assembly and structural variant detection
To reduce library preparation time
To increase read accuracy
Explanation - Mate‑pair reads are generated from DNA fragments with large insert sizes, aiding scaffolding.
Correct answer is: To span large insert sizes for assembly and structural variant detection
Q.170 Which of the following best explains 'adapter contamination' in sequencing data?
The presence of short adapter sequences in reads that can interfere with alignment
The presence of high‑quality reads that are not properly trimmed
The presence of long reads that exceed the expected length
The presence of duplicated reads from PCR amplification
Explanation - Adapter contamination can cause misalignment and false variant calling if not trimmed.
Correct answer is: The presence of short adapter sequences in reads that can interfere with alignment
Q.171 In Illumina sequencing, what does the 'cluster density' affect?
Read length
Sequencing error rate and throughput
Base‑calling speed
Quality of adapters
Explanation - Too high cluster density can cause signal overlap and errors, while too low density reduces throughput.
Correct answer is: Sequencing error rate and throughput
Q.172 Which of the following sequencing technologies is best suited for generating ultra‑long reads for telomere sequencing?
Illumina NovaSeq
PacBio Sequel IIe
Oxford Nanopore PromethION
Ion Torrent Proton
Explanation - PromethION can produce reads over 100 kb, ideal for resolving telomeric repeats.
Correct answer is: Oxford Nanopore PromethION
Q.173 Which of the following best describes the role of 'phasing' in Illumina sequencing?
The process of aligning reads to a reference genome
The synchronization of nucleotide incorporation across clusters during sequencing
The detection of indels in homopolymer regions
The removal of adapter sequences
Explanation - Phasing refers to mis‑synchronization of base incorporation among DNA strands in a cluster.
Correct answer is: The synchronization of nucleotide incorporation across clusters during sequencing
Q.174 Which of the following best explains a 'coverage uniformity' metric?
The consistency of read length across a dataset
The consistency of sequencing depth across genomic regions
The consistency of adapter contamination across samples
The consistency of base quality scores across reads
Explanation - Uniform coverage ensures no regions of the genome are under‑represented.
Correct answer is: The consistency of sequencing depth across genomic regions
Q.175 Which of the following sequencing platforms uses 'laser‑excited fluorescence' for base detection?
Illumina HiSeq
PacBio Sequel II
Oxford Nanopore MinION
Ion Torrent PGM
Explanation - Illumina’s sequencing‑by‑synthesis relies on laser‑excited fluorescent signals.
Correct answer is: Illumina HiSeq
Q.176 Which of the following best describes 'single‑end sequencing'?
Sequencing reads from both ends of a DNA fragment
Sequencing one end of a DNA fragment only
Sequencing an entire genome in one run
Sequencing only the coding regions of a genome
Explanation - Single‑end sequencing generates a read from a single end of each DNA fragment, unlike paired‑end.
Correct answer is: Sequencing one end of a DNA fragment only
Q.177 Which of the following is NOT a step in Illumina library prep?
Fragmentation
End‑repair and A‑tailing
Adapter ligation
Polymerase chain reaction amplification
Explanation - While PCR can be used in Illumina library prep, a PCR‑free method is also common; the question asks for a step that is not part of the standard pipeline.
Correct answer is: Polymerase chain reaction amplification
Q.178 What is a 'base‑calling error' in sequencing?
A mismatch between the sequenced base and the reference genome
Misidentification of the nucleotide during base calling
A missing base due to sequencing failure
A duplicate read caused by PCR amplification
Explanation - Base‑calling error occurs when the sequencer incorrectly assigns a nucleotide based on the detected signal.
Correct answer is: Misidentification of the nucleotide during base calling
Q.179 Which sequencing technology can sequence native RNA without reverse transcription?
Illumina sequencing
PacBio SMRT
Oxford Nanopore Direct RNA
Ion Torrent sequencing
Explanation - Nanopore can sequence RNA directly, preserving RNA modifications.
Correct answer is: Oxford Nanopore Direct RNA
Q.180 Which of the following best explains why Illumina sequencing can suffer from 'phasing errors'?
The fluorescence signal decays over time
Some DNA strands incorporate nucleotides at slightly different rates across cycles
The polymerase stalls during sequencing
The flow cell surface degrades during a run
Explanation - Phasing errors arise when strands in a cluster fall out of sync, leading to signal mixing.
Correct answer is: Some DNA strands incorporate nucleotides at slightly different rates across cycles