Molecular Diagnostics Exam 1 and 2

DNA fragments that are 8, 12 and 15 kb in size need to be resolved on a gel. Of the following percent agarose gels, which would be best to use for this purpose? 2.0% 3.0% 0.7% 1% 1.5%

0.7%

Which of the following is the best range for a primer length for a PCR reaction designed to amplify a specific product? 50-100 bp 10-100 bp 5-10 pb 15-30 bp 1-20 bp

15-30 bp

How many total autosomes do human cells have? 46 total 22 total 23 total 23 pairs 22 pairs 45 total

22 pairs

For the following DNA sequence, what would the Tm be for the molecule? GGG AGT GCG TAT 34°C 60°C 38°C 42°C

38C

For the following DNA sequence, what would the Tm be for the molecule? GGG AGT GCG TAT

38C

Which of the following is an example of recombinant DNA? A bacterial plasmid containing a piece of human DNA Bacterial DNA cut into a linear piece of DNA None of these A viral genome replicating within a bacterial cell Human genomic DNA digested with a type II restriction enzyme

A bacterial plasmid containing a piece of human DNA

The LOL gene spans 1 kb and contains 9 exons. Each exon encodes 25 amino acids. Within this gene sequence, which type of mutation will most likely cause the greatest phenotypic change (i.e. change the function of the gene product)? A frameshift mutation in the third codon of exon 1 A non-conservative mutation in the last codon in exon 9 A silent mutation in the middle of exon 2 A silent mutation in intron 1 of the gene A conservative mutation in the last exon (exon 9) of the gene sequence

A frameshift mutation in the third codon of exon 1

Identify two assay types that could be used for the detection of a single base pair mutation in a gene but that do NOT use PCR amplification as a part of the procedure. Briefly explain how the mutation is detected.

A microarray hybridization and ASO methods can detect a single nucleotide mismatch due to its high stringency requirement. This is due to the binding of the probe being blocked from the mismatch

A point mutation can have one of three effects on a gene protein coding region. List the 3 options and briefly describe what each mutation causes in the gene product. (mutation—effect)

A point mutation can cause: Missense: change in amino acid Nonsense: premature stop, formation of a stop codon Silent: No change/effect in amino acid

For enrichment of mRNA in an RNA extraction, which of the following steps would be added to the protocol? a second precipitation with isopropanol a step to precipitate the sample in chelex resin a step adding a polyT column to the sample prep a step to remove proteins with a phenol:chloroform extraction a step to add an additional alcohol precipitation

A step adding a polyT column to the sample prep

Which of the following is a key advantage of capillary gel electrophoresis over slab gel electrophoresis? B. Uses a gel matrix C. Separates based on size A. Requires smaller amount of sample. D. both C and B of these E. A, B and C of these

A. Requires smaller amount of sample

What type of staining is used in a Reverse banding karyotype? Ethidium bromide Giemsa SyBr Green Crystal violet Acridine orange

Acridine orange

The dot blot method is used in which one of the following mutation detection assays? Allele specific oligomer (ASO) hybridization Single strand conformation polymorphism (SSCP) Microarray analysis Melt curve analysis Denaturing gradiet gel electrophoresis (DGGE)

Allele specific oligomer (ASO) hybridization

In what phase of meiosis do sister chromatids chromosomes separate? Anaphase II Interphase Metaphase I Prophase II Metaphase II Prophase I Anaphase I

Anaphase II

23andMe is a private company that provides direct to consumer genetic information. Currently, they provide genetic information on which of the following to the consumer? Ancestry Complete DNA sequence Ancestry, carrier status for some inherited diseases, and complete DNA sequence information. Carrier status for some inherited diseases Ancestry and Carrier status for some inherited diseases.

Ancestry and carrier status

For a double-stranded DNA sample, the A260 is 0.7, and the total amount of sample is 0.5 ml. What is its concentration of the sample in ug/ml, and what is the total yield of DNA in the solution in ug? Show your work (i.e. formula) and steps in calculating.

Concentration = conversion factor (50) x A260 = 50 x .7 = 35microgram/mL Yield= concentration x mL of sample = 35 x .5 = 17.5micrograms/mL

Of the following assays, which works most similarly to how a Taqman probe works in a qPCR assay to generate a signal? Ligase chain reaction QB replicase Cycling probe Hybrid Capture bDNA

Cycling probe

Which of the following is a component of the nucleosome? D. A, B and C of these C. H3 A. Histone H2A E. only A and C of these B. DNA

D. A, B and C of these

To prevent primer-dimer formation, which strategy below would you apply? Design primer with a low Tm B) Design primers with a high Tm Design primers so that there are no interstrand homologies Design primers such that there is a 10°C difference between them in the Tm Design primers so that there are no intrastrand homologies

Design primers so that there are no interstrand homologies

Which of the following can result in a Restriction Fragment Length Polymorphism? B. a base pair change in the DNA that destroys a RE recognition site D. both A and B of these A. a base pair change in the DNA that creates a RE recognition site C. a change in the number of nucleotides between two RE recognition sites E. A, B and C of these

E. A, B and C of these

Which of the following conditions affect the migration of nucleic acid fragments on an agarose gel? A. gel percentage C. gel temperature E. A, B and C of these D. A and B of these B. voltage applied

E. A, B and C of these

Of the following techniques, which can be used to assess a patient sample for unknown mutations in within a specific gene? D. A or B of these A. Single Strand Conformation Polymorphism (SSCP) analysis C. Denaturing gradient gel electrophoresis (DGGE) analysis E. A, B or C of these B. Heteroduplex analysis

E. A, B or C of these

Watson and Crick used the research of others to solve the structure of DNA. Of the following, which was contributed by Rosalind Franklin's X-ray diffraction data? D. Both A and B these B. DNA was helical E. Both B and C of these A. The amount of adenines and pyrimidines is constant for a species C. The width of the DNA structure was uniform

E. both B and C of these

In which type of qPCR detection method is the probe in a secondary structure prior to hybridization to the target DNA? D. Molecular beacons A. Taqman probes B. Scorpion probes F. Both B and D of these E. Both A and C of these C. FRET probes

F. Both B and D of these

Which type of fluorescent detection probe system require two probes that are immediately adjacent on the target DNA template? Taqman probes Molecular Beacon probes Scorpion probes Scorpion probes FRET probes

FRET probes

Based on the electropherogram shown, what is the sex of this individual? XXY XYY Female Male

Female

Which of the following methods could be used to determine if a patient was aneuploid for the entire chromosome 18? D. karyotype F. B or C of these E. B, C or D of these G. A, B C or D of these B. metaphase FISH C. interphase FISH A. CGH

G. A, B C or D of these

What type of staining is used in the traditional karyotype? SyBr Green Acridine orange Giemsa Ethidium bromide Crystal violet

Giemsa

In which of the following assays are mutations detected by resolving short double-stranded DNA fragments on a non-denaturing gel in which there can be a single base pair mismatch if a mutation is present? Heteroduplex analysis AS-PCR ASO Southern blot SSCP

Heteroduplex analysis

Other than sequencing, identify a method to detect a gene mutation biochemically and briefly describe how this method detects the mutation (1-2 sentences).

Immunoassay methods can detect a gene mutation by targeting a specific protein such as an antibody or antigen. Immunoassay can include enzymes that detect directly, indirectly, or capture the antigen. Mutations can be detected by the results of binding. For example with enzyme assays, the enzyme present is proportional to the amount of protein present. If the targeted protein is not present, the enzyme will not be detected.

The length of which phase, or which phase transition, in a qPCR reaction is the point that allows us to quantify the starting template DNA in the reaction? Exponential --> linear phase Plateau phase Linear phase Lag phase Linear --> plateau phase Exponential phase

Lag phase

Which of the following is NOT a denaturing agent? Urea formamide SDS MOPS

MOPS

Of the following, in the PCR reaction which would be the least likely to inhibit the reaction? EDTA SDS MgCl2 phenol urine

MgCl2

Using an amino acid coding chart, and given the mRNA sequence below, which of the following peptide sequences would be synthesized? mRNA sequence: AUGCAGCAUCGC

NH2-Met-Gln-His-Arg-COOH

In a research lab, Y chromosome DNA is being haplotyped and compared across populations throughout Africa and the Caribbean. Of the following, what is the most likely focus of the researcher's study? Criminal suspect identification Maternal lineage Assay development Paternal lineage Forensic analysis

Paternal linkage

Which of the following reagents can be used in a PCR reaction, and with UV light, controls for potential contamination of the reaction with previously amplified amplicon? UNG Urea exonuclease Glycerol Psoralen

Psoralen

For which type of electrophoresis should DNA samples be prepared within an agarose gel plug? pulsed field gel electrophoresis capillary Southern agarose polyacrylamide

Pulse field gel electrophoresis

Of the following, which method is used for probe amplification? Qβ replicase Hybrid capture bDNA Polymerase chain reaction Cycling probe TMA

QB replicase

Which of the following is a type of probe amplification system that uses magnetic beads? Strand displacement amplification QB replicase Transcription-based amplification Hybrid Capture Ligase chain reaction

QB replicase

Which of the following is NOT a characteristic of restriction enzymes (RE)? Each RE recognizes a specific double-stranded DNA binding site The normal function of a RE is to protect the cell from invading DNA RE can ligate two DNA strands together RE are bacterial enzymes

RE can ligate two DNA strands together

SHORT ANSWER: ANSWER IN 2-3 SENTENCES List 3 ways that RFLPs are used in clinical assays.

RFLPs are used in clinical assays by cleaving the target DNA sequences. They are used to detect mutations, and can be used for paternity testing and genotyping. RFLPS can be used to detect hereditary diseases as well.

Of the following, which pulsed field gel electrophoresis system has the electric poles in a fixed position with no alternating positions of the poles? TAFE CHEF Isoelectric focusing gel RGE SDS-PAGE

RGE

A Zoo blot is a membrane which contains samples of which of the following type of molecules? DNA RNA DNA or RNA Protein DNA or protein

RNA

List two differences between DNA isolation versus RNA isolation using organic methods. Briefly explain why these differences exist. (Answer as difference - reason).

RNA is less stable than DNA therefore we need to limit the number of RNases present. We can do this by performing the isolation experiment in and RNase free zone and use RNase free equipment. RNA is less stable due to the ribose structure. We use a detergent that inhibits the RNases. DEPC TREATMENT In DNA organic isolation we use substances like SDS or NaOH to lyse the cells but use a detergent in RNA isolation that acts as the RNase inhibitor. In DNA extraction we also have to acidify the solution but do not need to in RNA isolation because it is already in an acid solution.

Which modification of PCR uses RNA-directed DNA polymerase? multiplex PCR RAPD PCR nested PCR RT-PCR qPCR

RT=PCR

Of the following, which PCR control would best reveal contamination of the reaction with target DNA? Negative template strand control Reagent blank, Amplification control, and Negative template strand control Amplification control Reagent blank Reagent blank and Negative template strand control

Reagent blank and Negative template strand control

The product of crossing over events in meiosis are _______. Sister chromatids Asters Recombinant chromosomes Parental chromosomes Haploid cells

Recombinant chromosomes

Which of the following statements is an accurate statement comparing replication to transcription? Replication uses a DNA polymerase in the nucleus while transcription uses an RNA polymerase in the cytoplasm and Replication uses a DNA polymerase while transcription uses RNA polymerase with both occurring in the nucleus Transcription uses a DNA polymerase in the nucleus while replication uses an RNA polymerase in the cytoplasm. Replication uses a DNA polymerase in the nucleus while transcription uses an RNA polymerase in the cytoplasm. Replication uses a DNA polymerase while transcription uses RNA polymerase with both occurring in the nucleus.

Replication uses a DNA polymerase while transcription uses RNA polymerase with both occurring in the nucleus.

Which of the following is NOT a basic component of a standard PCR reaction? primers KCl dNTPs polymerase reverse transcriptase

Reverse transcriptase

In which method of gene mutation analysis is only single-stranded DNA fragments resolved on a gel? Melt curve analysis ASO hybridization SSCP Heteroduplex analysis DGGE

SSCP

The majority of markers used in the CODIS system are what type of polymorphic marker sequences? SNP RFLP VNTR STR LINES

STR

In what type of procedure is neither the target nor probe immobilized onto a membrane? Chip array Dot blot Southern blot Microarray Solution hybridizaiton

Solution hybridization

Of the following, which solution is commonly used to resuspend a DNA sample in at the end of an isolation / purification protocol? SDS (Sodium dodecyl sulfate) CHAPS (3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate) EDTA (Ethylenediaminetetraacetic acid) TE (Tris EDTA) Isopropanol

TE (Tris EDTA)

Which of the following is added to an acrylamide - BIS solution to stimulate polymerization? (check all that apply) dextran TEMED MOPS APS SDS CHAPS

TEMED, APS

Which of the following qPCR detection probes relies on an exonuclease function contained within a polymerase to degrade the probe to release the Reporter dye from proximity to the quencher dye? Taqman probes Both Molecular beacon and Scorpion probes FRET probes Scorpion probes Molecular beacons

Taqman probes

In your gel electrophoresis, the migration of the samples, including the marker, is progressing down the gel more quickly than expected, i.e. the samples are migrating more quickly than they should. Identify two things that could be contributing to this issue, and for each, explain how each could contribute to the issue.

The %gel matrix may be too low allowing for larger pores for the samples to migrate through quickly. Another thing that could contribute to this issue is using too high of a voltage. The voltage is what migrates the samples through the buffer.

Melt curve analysis of a mutation is based on the sequence changes affecting Tm primer binding amplification amount of DNA present

Tm

For the analysis of PCR products, dUTP and UNG are used for what purpose? To prevent contamination with amplicon from previous reactions To increase the specificity of the reaction To prevent mispriming To prevent primer dimer formation To prevent bacterial DNA contamination

To prevent contamination with amplicon from previous reactions

The following is a list of characteristics of a particular method of nucleic acid amplification. Choose the method that is best described by these characteristics: Requires two primers, one is a tailed primer, and one is a standard primer The tailed primer contains an RNA polymerase promoter Uses three enzymes in the reaction (reverse transcriptase, an RNase & RNA polymerase ) Is an isothermal reaction Product is an RNA transcript: Strand Displacement Amplification (SDA) Transcription-based Amplification system (TAS) branched DNA (bDNA) hybrid capture Qbeta replicase Ligase Chain Reaction (LCR)

Transcription-based Amplification system (TAS)

Choose either the Transcription-based amplification syste (TAS) or the Strand Displacement Assay (SDA) assay. First write which assay you chose, then briefly identify the necessary reagents required for assay, and a short description of how the assay works to amplify the nucleic acid.

Transcription-based amplification requires enzymes RNase, RNA transcriptase and RNA polymerase. The target sequence is RNA and is amplified by having a DNA intermediate. Both a tailed RNA polymerase promoter and a standard primer are used. A DNA intermediate is formed off of the original RNA sequence. The cDNA is then used to transcribe RNA. Multiple cycles are performed to amplify the RNA target.

Of the following amplification methods, which uses RNA polymerase in the reaction? Ligase chain reaction (LCR) qPCR Transcription-based amplification system (TAS) Strand Displacement Amplification (SDA) nested PCR

Transcription-based amplification system (TAS)

List two factors that contribute to the effective Tm of a probe hybridization, and briefly describe how increasing the factor in a hybridization would affect the Tm. (Answer as factor - affect).

Two factors that contribute to the effect of Tm of a probe are salt concentration and denaturing agents. An increase in salt concentration will increase Tm because salt increases the melting point and an increase in denaturing agent will decrease the Tm because the denaturing agent affects the hydrogen bonds.

Of the following, which mechanism of epigenetic gene regulation utilizes chromatin condensation? Imprinting Transcriptional inhibitor binding Gene promoter methylation Transcriptional enhancer binding X-inactivation

X inactivation

Putrefaction of a human tissue or cell sample is caused by which of the following? contamination of sample with exogenous DNA drying out of the sample due to removal from host none of these autolysis of the sample due to removal from host infection of the sample with bacteria

autolysis of the sample due to removal from host

Capture probes, extender probes and amplifiers are all used in which of the following reactions? Cycling probe Ligase chain reaction (LCR) Strand displacement amplification (SDA) branched DNA (bDNA) Cleavage based amplification

branched DNA (bDNA)

Which of the following assays requires an extender probe in the reaction? Ligase chain reaction (LCR) QB replicase branched DNA (bDNA) Hybrid capture Cycling probe

branched DNA (bDNA)

Which of the following types of probe detection develops color on the blot itself and thus only allows one exposure to be obtained? chemifluorescent radioactive chemiluminescent chromogenic

chromogenic

A karyotype that shows a patient to be 46, XX would be most appropriately described as ____. diploid monoploid aneuploid polyploid haploid

diploid`

STRs and SNPs are often used in genetic analysis. Some STRs and SNPs are within genes, but most are not. Of the following applications, which would use non-gene STRs and/or SNPs? Forensics, Paternity tests, and Hereditary disease diagnosis Paternity tests Forensics and Paternity tests Hereditary disease diagnostics Forensics

forensics and paternity tests

Which of the following chemicals is used to break DNA strands at the G or an A nucleotide for sequencing reactions? Hydrazine Dimethylsulfate Formic acid Hydrazine + salt Piperidine

formic acid

A series of polymorphism alleles, along the same segment of a chromosome locus, that are inherited together as a unit are called a _______. BLAST polymorphism mutation homolog haplotype

haplotype

Which of the following types of RNA is a precursor to mRNA in eukaryotes? tRNA miRNA hnRNA rRNA siRNA

hnRNA

In Maxam-Gilbert sequencing, which chemical method is used to break DNA chains at cytosine residues? dimethylsulfate hydrazine + salt urea hydrazine formic acid

hydrazine and salt

For a double stranded oligonucleotide, increasing the G and C nucleotide content would ______ the Tm while decreasing the length of the nucleotide would ______ the Tm? decrease, decrease decrease, increase increase, decrease increase, increase cannot be determined with this information

increase, decrease

Which of the following is the correct order of basic steps in a nucleic acid extraction? pellet debris → lyse cells → separate cellular molecules (phase separation) → alcohol precipitate DNA separate cellular molecules (phase separation) → lyse cells → alcohol precipitate DNA → pellet debris lyse cells → pellet debris → separate cellular molecules (phase separation) → alcohol precipitate DNA lyse cells → separate cellular molecules (phase separation) → pellet debris → alcohol precipitate DNA

lyse cells → pellet debris → separate cellular molecules (phase separation) → alcohol precipitate DNA

Within an animal cell, a protein becomes damaged and nonfunctional. Which of the following organelles could be responsible for removing this damaged protein form the cell? Mitochondria Golgi apparatus Lysosome Endoplasmic reticulum Nucleus

lysosome

In the first step of translation initiation, what 3 things come together first? mRNA, large ribosomal subunit, small ribosomal subunit mRNA, splicesome, small ribosomal subunit mRNA, large ribosomal subunit, initiator tRNA mRNA, small ribosomal subunit, initiator tRNA

mRNA, small ribosomal subunit, initiator tRNA

Of the following, which sequencing methods uses chemical breakdown of the DNA to obtain sequence information? Fourth generation sequencing Maxam-Gilbert Third generation sequencing Second generation sequencing Sanger dideoxy sequencing

maxam-gilbert

At what stage of the cell cycle is a standard karyotype stained? telophase interphase prophase anaphase metaphase

metaphase

When a set of primers designed to amply the target region, A, also amplifies another region of the organism's genome at a larger bp size, this is called a ______. primer dimer intrastrand homology a bonus product interstrand homology misprime

misprime

Which of the following structures is only in a eukaryotic cell? plasma membrane mitochondria DNA polymerases ribosome

mitochondria

Of the following, which is NOT a type of repeating DNA segment sequence? VNTR neither STR and SNP SINES STR SNP HLA locus neither SNP and HLA locus neither STR, VNTR, SNP.

neither SNP or HLA locus

What type of PCR reaction uses two sets of primers sequentially, with the second set of primers contained within the amplicon of the first set of primers? RT-PCR multiplex PCR qPCR nested PCR RAPD PCR

nested PCR

A mutation occurs within the coding sequence of a gene. This mutation substitutes one nucleotide for another which results in the introduction of a stop codon. What type of mutation is this? frameshift missense sense nonsense silent

nonsense

Of the following blotting methods, which have patient sample (DNA or RNA) fixed to the solid support? Northern blot and dot blot macroarray dot blot Northern blot Microarray and macroarray microarray Microarray and dot blot

northern blot and dot blot

While DNA can be extracted from saliva, for analysis of a DNA segment of interest, it will most likely require which of the following procedures? both organic and inorganic extractions methods nucleic acid amplification (g. polymerase chain reaction) of the target DNA multiple alcohol precipitations skipping the cell lysis step a nuclease treatment step

nucleic acid amplification (PCR) of the target DNA

For a sequencing application, you need a DNA sample of high purity. When you calculate the absorbance ratio at A260/A280 for your sample, it is 1.4. Of the following, which is the best way to proceed? re-precipitate the DNA in ethanol throw the sample out perform another phenol:chloroform extraction on the sample tell your boss it wasn't your fault Use the sample as it is

perform another phenol:chloroform extraction on the sample

What structure in conjugation connects the two bacterial cells? free double stranded DNA flagella drug resistant plasmid pilus bacteriophage

pilus

You are preparing to run a gel and have dissolved your agarose in buffer in the microwave when you realize that you forgot to prepare the casting tray. Of the following options, which would be the best option to proceed? place the agarose solution in a 55°C water bath place the agarose solution in the refrigerator pour the agarose solution down the drain and start over place the agarose solution in a 40°C water bath leave the agarose solution on the bench top until the tray is ready

place the agarose solution in a 55°C water bath

A diploid cell is fused haploid cell in a cell culture experiment. The resulting cell would best be described as ____. haploid aneuploid diploid Polyploid monoploid

polyploid

What enzyme synthesizes an RNA primer during DNA replication in prokaryotes? topoisomerase DNA polymerase I primase nuclease DNA polymerase alpha

primase

Within a replication fork, which of the following enzymes in required only one time for the leading strand, but multiple times for the lagging strand? primase helicase exonuclease DNA polymerase RNA polymerase

primase

During what phase of the cell cycle do replicated chromosomes condense? telophase prophase metaphase G1 phase S phase

prophase

Which type of sequencing reactions uses luciferase activity to generate signal for detection? Maxam-Gilbert Sanger chain termination Bisulfite Pyrosequencing Pyrosequencing and Bisulfite sequencing

pyrosequencing

The long arm of an acrocentric chromosome is designated as which of the following? S arm telomere p arm q arm L arm

q arm

In which type of assay is an antibody fixed on a solid surface to which an antigen is bound followed by detection with a second, labeled antibody? AS-PCR SSCP sandwich ELISA HPLC indirect ELISA

sandwich ELISA

DNA is to the _____ blot as RNA is to the _____ blot. Western, Southern Southern, Southwestern Southern, Northern Southern, Western Northern, Southern

southern, northern

The GGU codon encodes only for the amino acid glycine (Gly). What principle of the genetic code does this represent? wobble ambiguity universality specificity redundancy

specificity

Of the following types of RNA, which functions in translation to bring amino acids to the ribosome? mRNA snRNA hnRNA tRNA rRNA

tRNA

In a qPCR reaction, the length of the lag phase, or time to threshold, is inversely proportional to the concentration of ________ in the reaction. none of these primers template DNA fluorescent probe thermostable DNA polymerase

template DNA