DNA SEQUENCING

*Question: What is DNA sequencing?
A) A method to amplify DNA fragments
B) A technique to determine the exact sequence of nucleotides in a DNA molecule
C) A process to visualize DNA using gel electrophoresis
D) A method to extract DNA from cells

*Answer: B) A technique to determine the exact sequence of nucleotides in a DNA molecule

*Question: Which letters represent the four nitrogenous bases in DNA?
A) A, U, C, G
B) A, T, C, G
C) A, T, C, U
D) A, T, G, U

*Answer: B) A, T, C, G

*Question: DNA sequencing is considered the gold standard for diagnosing which of the following?
A) Bacterial infections only
B) Viral infections only
C) Genetic disorders, cancer mutations, infectious agents, and inherited diseases
D) Autoimmune disorders only

*Answer: C) Genetic disorders, cancer mutations, infectious agents, and inherited diseases

*Question: What is the most definitive molecular method to identify genetic lesions?
A) PCR
B) Gel electrophoresis
C) DNA sequencing
D) Hybridization

*Answer: C) DNA sequencing

*Question: In the molecular biology workflow, what comes after gel electrophoresis?
A) PCR amplification
B) DNA extraction
C) Other downstream applications
D) Nucleic acid quantification

*Answer: C) Other downstream applications

*Question: Why is it important to check the quality and purity of DNA before PCR?
A) To determine the sequence of the DNA
B) To avoid wasting resources on degraded or contaminated samples
C) To identify which gene to target
D) To visualize the DNA fragments

*Answer: B) To avoid wasting resources on degraded or contaminated samples

*Question: What does gel electrophoresis confirm after PCR?
A) The sequence of the DNA
B) The purity of the DNA
C) Whether the target gene was amplified
D) The quantity of the DNA

*Answer: C) Whether the target gene was amplified

*Question: Which tool allows monitoring of amplification in real time due to fluorescent dye?
A) Conventional PCR
B) Gel electrophoresis
C) Real-time PCR
D) Capillary electrophoresis

*Answer: C) Real-time PCR

*Question: Who pioneered DNA sequencing in 1977?
A) Watson, Crick, and Franklin
B) Sanger, Maxam, and Gilbert
C) Mullis, Sanger, and Watson
D) Gilbert, Crick, and Mullis

*Answer: B) Sanger, Maxam, and Gilbert

*Question: Which sequencing method was published first in 1977?
A) Sanger method
B) Capillary sequencing
C) Maxam-Gilbert method
D) Pyrosequencing

*Answer: C) Maxam-Gilbert method

*Question: What month and year did Sanger publish his sequencing method?
A) February 1977
B) June 1977
C) December 1977
D) March 1977

*Answer: C) December 1977

*Question: What type of method does Maxam-Gilbert sequencing use?
A) Synthesis-based method
B) Chemical degradation method
C) Fluorescence-based method
D) Electrical current method

*Answer: B) Chemical degradation method

*Question: Which of the following are First Generation Sequencing technologies?
A) Illumina, Pyrosequencing, PacBio
B) Maxam-Gilbert, Sanger, Capillary Sequencing
C) PacBio, Oxford Nanopore, Illumina
D) Sanger, Illumina, Oxford Nanopore

*Answer: B) Maxam-Gilbert, Sanger, Capillary Sequencing

*Question: What principle does the Capillary Sequencing (CapSeq) method use?
A) Chemical degradation
B) Electrical current changes
C) Sanger technique
D) Bridge amplification

*Answer: C) Sanger technique

*Question: In Maxam-Gilbert sequencing, how many reaction tubes is the DNA split into?
A) 2
B) 3
C) 4
D) 5

*Answer: C) 4

*Question: What is the role of piperidine in Maxam-Gilbert sequencing?
A) It labels the DNA radioactively
B) It initially alters the structure of bases
C) It cleaves or breaks the DNA fragments
D) It separates fragments by size

*Answer: C) It cleaves or breaks the DNA fragments

*Question: Which chemical is used in Maxam-Gilbert sequencing to target guanine?
A) Formic acid
B) Hydrazine
C) Hydrazine + salt
D) Dimethylsulphate

*Answer: D) Dimethylsulphate

*Question: Which chemical targets purines (G and A) in Maxam-Gilbert sequencing?
A) Hydrazine
B) Formic acid
C) Dimethylsulphate
D) Hydrazine + salt

*Answer: B) Formic acid

*Question: Which chemical targets pyrimidines (C and T) in Maxam-Gilbert sequencing?
A) Formic acid
B) Dimethylsulphate
C) Hydrazine
D) Piperidine

*Answer: C) Hydrazine

*Question: Which chemical specifically targets cytosine only in Maxam-Gilbert sequencing?
A) Hydrazine
B) Formic acid
C) Dimethylsulphate
D) Hydrazine + salt

*Answer: D) Hydrazine + salt

*Question: How is piperidine removed after cleavage in Maxam-Gilbert sequencing?
A) Centrifugation
B) Gel electrophoresis
C) Evaporation through repeated drying and alcohol washes
D) Filtration

*Answer: C) Evaporation through repeated drying and alcohol washes

*Question: What solution is used to resuspend the DNA after drying in Maxam-Gilbert sequencing?
A) Ethanol
B) Formamide
C) Piperidine
D) Formic acid

*Answer: B) Formamide

*Question: Why is polyacrylamide gel preferred over agarose gel in Maxam-Gilbert sequencing?
A) It is cheaper and easier to use
B) It is suitable for fragments less than 500 bp and has high resolving power
C) It does not require radioactive labeling
D) It is compatible with fluorescent dyes

*Answer: B) It is suitable for fragments less than 500 bp and has high resolving power

*Question: In Maxam-Gilbert sequencing, which end of the DNA is radioactively labeled?
A) 3' end
B) Middle of the strand
C) 5' end
D) Both ends

*Answer: C) 5' end

*Question: Which strand is preferred for sequencing in Maxam-Gilbert?
A) Antisense strand (3' to 5')
B) Sense strand or nontemplate strand (5' to 3')
C) Template strand (3' to 5')
D) Either strand

*Answer: B) Sense strand or nontemplate strand (5' to 3')

*Question: How are DNA fragments read in gel electrophoresis after Maxam-Gilbert sequencing?
A) Top to bottom (nearest to farthest from well)
B) From the middle outward
C) Bottom to top (farthest to nearest from well)
D) Left to right

*Answer: C) Bottom to top (farthest to nearest from well)

*Question: In gel electrophoresis, which fragment migrates the farthest from the well?
A) The longest fragment
B) The fragment with the most bases
C) The shortest fragment
D) The heaviest fragment

*Answer: C) The shortest fragment

*Question: In Maxam-Gilbert gel reading, if a band appears in both the 3rd lane (C+T) and 4th lane (C only), what base is it?
A) Thymine
B) Guanine
C) Adenine
D) Cytosine

*Answer: D) Cytosine

*Question: In Maxam-Gilbert gel reading, if a band appears in the 3rd lane (C+T) but NOT in the 4th lane (C only), what base is it?
A) Cytosine
B) Guanine
C) Thymine
D) Adenine

*Answer: C) Thymine

*Question: In Maxam-Gilbert gel reading, if a band appears in the 2nd lane (G+A) but NOT in the 1st lane (G only), what base is it?
A) Guanine
B) Cytosine
C) Thymine
D) Adenine

*Answer: D) Adenine

*Question: Why is the Maxam-Gilbert method no longer commonly used today?
A) It is too expensive
B) The chemicals used are toxic, combustible, irritant, and flammable
C) It cannot sequence double-stranded DNA
D) It requires too many tubes

*Answer: B) The chemicals used are toxic, combustible, irritant, and flammable

*Question: What is the hazard classification of dimethylsulphate used in Maxam-Gilbert?
A) Flammable
B) Irritant
C) Toxic and combustible
D) Corrosive

*Answer: C) Toxic and combustible

*Question: What is formic acid classified as in terms of safety?
A) Combustible
B) Irritant
C) Toxic
D) Flammable

*Answer: B) Irritant

*Question: What is hydrazine classified as in terms of safety?
A) Irritant
B) Combustible
C) Flammable
D) Toxic

*Answer: C) Flammable

*Question: What is the Sanger sequencing method also called?
A) Chemical degradation method
B) Bridge amplification method
C) Chain termination method
D) Sequencing by synthesis

*Answer: C) Chain termination method

*Question: Who developed the Sanger sequencing method and when?
A) Maxam and Gilbert, 1975
B) Frederick Sanger, 1977
C) Watson and Crick, 1953
D) Mullis, 1983

*Answer: B) Frederick Sanger, 1977

*Question: What is the key difference between a deoxynucleotide and a dideoxynucleotide?
A) Dideoxynucleotide lacks the phosphate group
B) Dideoxynucleotide has a hydrogen group at the 3' position instead of a hydroxyl group
C) Dideoxynucleotide has an extra oxygen molecule
D) Dideoxynucleotide lacks the nitrogenous base

*Answer: B) Dideoxynucleotide has a hydrogen group at the 3' position instead of a hydroxyl group

*Question: Why does chain termination occur when a dideoxynucleotide is incorporated?
A) It degrades the template strand
B) The polymerase enzyme detaches from the strand
C) There is no hydroxyl group at the 3' position for the next nucleotide to be added
D) It blocks the primer from binding

*Answer: C) There is no hydroxyl group at the 3' position for the next nucleotide to be added

*Question: In Sanger sequencing, which position of the dideoxynucleotide lacks an oxygen molecule?
A) 1' and 2' positions
B) Both 2' and 3' positions
C) 5' position only
D) 3' position only

*Answer: B) Both 2' and 3' positions

*Question: Which reagent is added in Sanger sequencing to promote equal or simultaneous addition of nucleotides?
A) Piperidine
B) Formamide
C) Manganese
D) Hydrazine

*Answer: C) Manganese

*Question: What does Sanger sequencing sequence, unlike Maxam-Gilbert?
A) The original sense strand
B) The newly synthesized complementary strand
C) The antisense strand
D) Double-stranded DNA directly

*Answer: B) The newly synthesized complementary strand

*Question: How many tubes are prepared in Sanger sequencing?
A) 2
B) 3
C) 5
D) 4

*Answer: D) 4

*Question: In Sanger sequencing, what does each tube contain?
A) A different chemical degradant
B) A different dideoxynucleoside triphosphate (ddNTP)
C) A different primer
D) A different restriction enzyme

*Answer: B) A different dideoxynucleoside triphosphate (ddNTP)

*Question: In Sanger sequencing, what happens to the base targeted by a dideoxynucleotide?
A) It is degraded and removed from the fragment
B) It is not included in the sequence
C) It is included in the fragment but terminates further chain elongation
D) It is replaced by another nucleotide

*Answer: C) It is included in the fragment but terminates further chain elongation

*Question: Which of the following is a key preparation component in Sanger sequencing NOT found in Maxam-Gilbert?
A) Radioactive label
B) DNA polymerase and dNTPs
C) Dideoxynucleoside triphosphate (ddNTP)
D) Polyacrylamide gel

*Answer: C) Dideoxynucleoside triphosphate (ddNTP)

*Question: In Sanger sequencing, from which tube does the shortest DNA fragment come?
A) ddATP tube
B) ddCTP tube
C) ddGTP tube
D) ddTTP tube

*Answer: D) ddTTP tube

*Question: What direction is the synthesized strand read in Sanger sequencing?
A) 3' to 5'
B) 5' to 3'
C) From the longest to the shortest
D) From the nearest to farthest from the well

*Answer: B) 5' to 3'

*Question: What is one advantage of Sanger sequencing over Maxam-Gilbert sequencing?
A) Uses cheaper reagents
B) Directly sequences each specific base since each tube is specific to one ddNTP
C) Does not require gel electrophoresis
D) Does not require radioactive labeling

*Answer: B) Directly sequences each specific base since each tube is specific to one ddNTP

*Question: What is the key difference between Capillary Sequencing (CapSeq) and Sanger sequencing?
A) CapSeq uses chemical degradation
B) CapSeq uses a narrow tube/capillary instead of gel and incorporates fluorescent dye in ddNTPs
C) CapSeq sequences the original sense strand
D) CapSeq uses piperidine for cleavage

*Answer: B) CapSeq uses a narrow tube/capillary instead of gel and incorporates fluorescent dye in ddNTPs

*Question: What advantage does CapSeq have over traditional Sanger sequencing?
A) It does not require primers
B) The fluorescent signal allows automatic detection of sequence order without manual gel reading
C) It sequences longer fragments
D) It is cheaper

*Answer: B) The fluorescent signal allows automatic detection of sequence order without manual gel reading

*Question: What are the two major categories of DNA sequencing methods?
A) Chemical and enzymatic sequencing
B) First generation and next generation sequencing
C) Manual and automated sequencing
D) Radioactive and fluorescent sequencing

*Answer: B) First generation and next generation sequencing

*Question: Which NGS type sequences the entire genome including both exon and intron regions?
A) Whole-exome sequencing
B) Targeted gene panel sequencing
C) Whole-genome sequencing
D) Amplicon sequencing

*Answer: C) Whole-genome sequencing

*Question: What percentage of the entire genome do exons represent?
A) 10–15%
B) 5–8%
C) 1–2%
D) 25–30%

*Answer: C) 1–2%

*Question: Which NGS type sequences only the protein-coding regions (exons)?
A) Whole-genome sequencing
B) RNA sequencing
C) Small RNA sequencing
D) Whole-exome sequencing

*Answer: D) Whole-exome sequencing

*Question: Which NGS type is best suited when you want to identify all organisms present in an environmental sample?
A) Whole-exome sequencing
B) Targeted gene panel sequencing
C) Amplicon sequencing
D) Metagenomic sequencing

*Answer: D) Metagenomic sequencing

*Question: What does RNA sequencing (RNA-seq) target?
A) All DNA molecules in the genome
B) Transcriptomes or all RNA molecules
C) Small non-coding RNAs only
D) Specific amplified regions

*Answer: B) Transcriptomes or all RNA molecules

*Question: What is the difference between Targeted Gene Panel Sequencing and Amplicon Sequencing?
A) TGPS targets the original strand while Amplicon Sequencing targets the amplified strand
B) TGPS uses fluorescence while Amplicon Sequencing uses radioactivity
C) TGPS is for RNA and Amplicon Sequencing is for DNA
D) They are identical methods

*Answer: A) TGPS targets the original strand while Amplicon Sequencing targets the amplified strand

*Question: What is the principle of pyrosequencing?
A) Electrical current changes identify each nucleotide
B) Light is produced only when the correct nucleotide is added, revealing the DNA sequence
C) Chemical degradation of bases produces fragments
D) Chain termination by dideoxynucleotides

*Answer: B) Light is produced only when the correct nucleotide is added, revealing the DNA sequence

*Question: What does SBS stand for in the context of sequencing?
A) Single Base Sequencing
B) Sequencing by Synthesis
C) Strand-Based Separation
D) Signal-Based Scanning

*Answer: B) Sequencing by Synthesis

*Question: Which sequencing method is pyrosequencing classified under?
A) 3rd generation
B) 1st generation
C) 4th generation
D) 2nd generation

*Answer: D) 2nd generation

*Question: Why was pyrosequencing discontinued?
A) It was too slow
B) It could not detect mutations
C) The chemical used is very toxic
D) It could not sequence RNA

*Answer: C) The chemical used is very toxic

*Question: What is pyrosequencing best used for?
A) Whole-genome sequencing
B) Novel sequence discovery
C) Short-read tasks like detecting SNPs, mutations, and DNA methylation
D) Metagenomic sequencing

*Answer: C) Short-read tasks like detecting SNPs, mutations, and DNA methylation

*Question: What is the reaction mix for pyrosequencing?
A) Template, primer, polymerase, dNTPs, ddNTPs
B) Template, primer, polymerase, nucleotide, APS, sulfurylase, luciferase, luciferin
C) Template, primer, polymerase, piperidine, formamide
D) Template, primer, ddNTPs, manganese

*Answer: B) Template, primer, polymerase, nucleotide, APS, sulfurylase, luciferase, luciferin

*Question: In pyrosequencing, what is released every time a nucleotide is added by the polymerase?
A) ATP
B) Luciferin
C) Pyrosulfate (PPi)
D) APS

*Answer: C) Pyrosulfate (PPi)

*Question: In pyrosequencing, what converts APS to ATP?
A) Luciferase
B) Polymerase
C) Sulfurylase (sulforase)
D) Luciferin

*Answer: C) Sulfurylase (sulforase)

*Question: In pyrosequencing, what converts luciferin into a light signal?
A) Sulfurylase
B) ATP (in the presence of luciferase)
C) Pyrosulfate
D) DNA polymerase

*Answer: B) ATP (in the presence of luciferase)

*Question: What is the principle of Illumina sequencing?
A) Chemical degradation of bases
B) Electrical current changes at nanopores
C) Massively parallel sequencing based on Sequencing by Synthesis (SBS)
D) Hairpin adapter-based long read sequencing

*Answer: C) Massively parallel sequencing based on Sequencing by Synthesis (SBS)

*Question: What generation is Illumina sequencing classified under?
A) 1st generation
B) 3rd generation
C) 4th generation
D) 2nd generation

*Answer: D) 2nd generation

*Question: Which sequencing method is the most commonly available in the Philippines?
A) PacBio
B) Oxford Nanopore
C) Illumina
D) Sanger

*Answer: C) Illumina

*Question: What are the five basic steps of Illumina sequencing?
A) Extraction, purification, amplification, hybridization, analysis
B) DNA/RNA fragmentation, library preparation, amplification, sequencing by synthesis, data analysis
C) Denaturation, annealing, extension, gel electrophoresis, analysis
D) Labeling, degradation, separation, detection, analysis

*Answer: B) DNA/RNA fragmentation, library preparation, amplification, sequencing by synthesis, data analysis

*Question: In Illumina sequencing, what is added to both ends of the fragmented DNA during library preparation?
A) Radioactive labels
B) Primers
C) Adapters
D) Dideoxynucleotides

*Answer: C) Adapters

*Question: What is the purpose of adapters in Illumina library preparation?
A) They degrade unwanted sequences
B) They serve as binding sites for primers and as labels for identification
C) They amplify the DNA fragments
D) They terminate chain elongation

*Answer: B) They serve as binding sites for primers and as labels for identification

*Question: In Illumina sequencing, what does denaturation accomplish during library preparation?
A) It removes adapters from the DNA
B) It fragments the DNA into smaller pieces
C) It separates the double-stranded DNA into single-stranded DNA
D) It amplifies the DNA

*Answer: C) It separates the double-stranded DNA into single-stranded DNA

*Question: What is the box-like structure used in Illumina amplification called?
A) PCR chamber
B) Flow well (flow cell)
C) Capillary tube
D) Nanopore membrane

*Answer: B) Flow well (flow cell)

*Question: What are the primers fixed to in the Illumina flow well called?
A) Floating primers
B) Adapter primers
C) Immobilized or stationary primers
D) Dideoxy primers

*Answer: C) Immobilized or stationary primers

*Question: Why are original DNA strands washed away after the first extension step in Illumina amplification?
A) They interfere with fluorescent signals
B) They are not fixed to the flow well, so they must be removed
C) They contain radioactive labels
D) They block the primers

*Answer: B) They are not fixed to the flow well, so they must be removed

*Question: What type of amplification occurs in Illumina sequencing when the synthesized strand connects to another available primer on the flow well?
A) Rolling circle amplification
B) Bridge amplification
C) Linear amplification
D) Isothermal amplification

*Answer: B) Bridge amplification

*Question: In Illumina sequencing, what color signal indicates Thymine was successfully incorporated?
A) Orange
B) Green
C) Red
D) Blue

*Answer: D) Blue

*Question: In Illumina sequencing, what color signal indicates Adenine was successfully incorporated?
A) Blue
B) Green
C) Orange
D) Red

*Answer: C) Orange

*Question: In Illumina sequencing, what color signal indicates Guanine was successfully incorporated?
A) Red
B) Blue
C) Orange
D) Green

*Answer: D) Green

*Question: In Illumina sequencing, what color signal indicates Cytosine was successfully incorporated?
A) Green
B) Orange
C) Red
D) Blue

*Answer: C) Red

*Question: In Illumina sequencing, which strands are washed away before the sequencing step?
A) Forward (red) strands
B) Adapter strands
C) Reverse (blue) strands
D) Template strands

*Answer: C) Reverse (blue) strands

*Question: What generation is PacBio sequencing?
A) 2nd generation
B) 1st generation
C) 4th generation
D) 3rd generation

*Answer: D) 3rd generation

*Question: What is the unique feature of the adapter used in PacBio sequencing?
A) It is radioactively labeled
B) It is a hairpin adapter
C) It is a chemical degradant
D) It is a fluorescent bead

*Answer: B) It is a hairpin adapter

*Question: What type of signal is used in PacBio sequencing to determine the DNA sequence?
A) Electrical current changes
B) Radioactive emissions
C) Chemical color changes
D) Fluorescence (light) signal upon correct nucleotide incorporation

*Answer: D) Fluorescence (light) signal upon correct nucleotide incorporation

*Question: What generation is Oxford Nanopore sequencing?
A) 2nd generation
B) 3rd generation
C) 1st generation
D) 4th generation

*Answer: D) 4th generation

*Question: What type of signal does Oxford Nanopore sequencing use to identify nucleotides?
A) Fluorescent light
B) Radioactive emissions
C) Electrical current changes
D) Color-coded signals

*Answer: C) Electrical current changes

*Question: Why does Oxford Nanopore sequencing deserve its own generation classification?
A) It sequences RNA instead of DNA
B) Its mechanism based on electrical current changes is fundamentally different from all prior methods
C) It uses the largest sequencing machines
D) It requires no sample preparation

*Answer: B) Its mechanism based on electrical current changes is fundamentally different from all prior methods

*Question: What structure does DNA pass through in Oxford Nanopore sequencing?
A) Capillary tubes
B) Flow wells
C) Tiny protein pores (nanopores) embedded in a membrane
D) Polyacrylamide gel

*Answer: C) Tiny protein pores (nanopores) embedded in a membrane

*Question: What is maintained constantly in Oxford Nanopore sequencing while DNA passes through the pore?
A) Temperature
B) Fluorescent signal
C) Electrical current
D) pH level

*Answer: C) Electrical current

*Question: How does Oxford Nanopore sequencing identify different nucleotides?
A) By the color of the fluorescent light emitted
B) By the weight of each fragment in gel electrophoresis
C) By the unique pattern change in electrical current as each nucleotide passes through the pore
D) By the length of synthesized fragments

*Answer: C) By the unique pattern change in electrical current as each nucleotide passes through the pore

*Question: What is the SMRTbell template in PacBio sequencing?
A) The flow well used for bridge amplification
B) The double-stranded DNA template with hairpin adapters on both ends
C) The nanopore membrane
D) The fluorescent dye incorporated into primers

*Answer: B) The double-stranded DNA template with hairpin adapters on both ends

*Question: Which sequencing method specifically CANNOT be used for whole-genome or novel sequence discovery?
A) Illumina
B) Oxford Nanopore
C) PacBio
D) Pyrosequencing

*Answer: D) Pyrosequencing

*Question: In Sanger sequencing, what does the DNA polymerase require to add a new nucleotide?
A) A free 5' phosphate group
B) A free 3' hydroxyl group
C) A dideoxynucleotide
D) A radioactive label

*Answer: B) A free 3' hydroxyl group

*Question: In Maxam-Gilbert sequencing with template ACG-TTG-GAC, using dimethylsulphate (G only), how many DNA fragments are produced?
A) 2
B) 4
C) 1
D) 3

*Answer: D) 3

*Question: What are the fragments produced when dimethylsulphate (G) is used on template ACG-TTG-GAC?
A) A, AC, ACG
B) AC, ACGTT, ACGTTG
C) A, ACGTT, ACGTTGGA
D) ACG, ACGTTG, ACGTTGGAC

*Answer: B) AC, ACGTT, ACGTTG

*Question: In Maxam-Gilbert sequencing with template ACG-TTG-GAC, what are the fragments when hydrazine + salt (C only) is used?
A) AC, ACGTT
B) ACG, ACGTTG
C) A, ACGTTGGA
D) AC, ACGTTGGA

*Answer: C) A, ACGTTGGA

*Question: In the example sequence ACG-TTG-GAC using Maxam-Gilbert, what is the shortest fragment?
A) AC
B) ACG
C) A
D) ACGTT

*Answer: C) A

*Question: Which method in first generation sequencing was first generation chemical-based?
A) Illumina and PacBio
B) Maxam-Gilbert and Sanger
C) CapSeq and Illumina
D) PacBio and Oxford Nanopore

*Answer: B) Maxam-Gilbert and Sanger

*Question: Why does sequencing only require one strand of DNA?
A) Two strands would produce too many fragments
B) The complementary strand can be inferred from base pairing rules
C) Polymerase only works on single-stranded DNA
D) Radioactive labeling only works on one strand

*Answer: B) The complementary strand can be inferred from base pairing rules