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Exam (elaborations)

FRNSC 421W - EXAM 1 PRACTICE QUESTIONS AND ANSWERS (100% PASS)

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  • FRNSC421W /FRNSC 420
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  • FRNSC421W /FRNSC 420

FRNSC 421W - EXAM 1 PRACTICE QUESTIONS AND ANSWERS (100% PASS) RFLP - Answer️️ -restriction fragment length polymorphism - DNA digested by restriction enzymes at repeat sequences, yields different fragment lengths for different people - Gel electrophoresis to separate fragments by size, MW...

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  • August 31, 2024
  • 30
  • 2024/2025
  • Exam (elaborations)
  • Questions & answers
  • FRNSC421W /FRNSC 420
  • FRNSC421W /FRNSC 420
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OliviaWest
©PREP4EXAMS2024/2025 REAL EXAMS DUMP Wednesday, August 7, 2024 9: 33 PM



FRNSC 421W - EXAM 1 PRACTICE QUESTIONS AND
ANSWERS (100% PASS)


RFLP - Answer✔️✔️-restriction fragment length polymorphism

- DNA digested by restriction enzymes at repeat sequences, yields different
fragment lengths for different people

- Gel electrophoresis to separate fragments by size, MW ladder for reference

- Southern blotting to physically transfer DNA to membrane

- Radioactively labeled probe hybridizes with immobilized DNA on membrane

- Exposure to X-ray film yields carbon copy of gel, reveals unique blotting pattern
specific to genotype

- Single locus probes use 1 probe to hybridize with multiple loci; only need 4-6
loci to support a match

- Labor intensive but highly discriminatory; requires high MW and high-quality
DNA

D1S80 - Answer✔️✔️-highly variable VNTR locus, separates DNA fragments by
allelic number (smaller fragments/allele # migrate further)

- Resolves alleles of loci and runs on a silver-stained gel; allelic ladder represents
known # of repeats

- DNA extraction and PCR amplification with silver staining instead of
radioactive/fluorescent probes

- can use on lower quality and amounts of DNA than RFLP



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,©PREP4EXAMS2024/2025 REAL EXAMS DUMP Wednesday, August 7, 2024 9: 33 PM


DQα - Answer✔️✔️-gene on chromosome 6 that codes for cell surface receptor
involved in immune response, SNP-based (PCR) reverse dot blot assay

- Immobilized probe fixed to nylon membrane by tails; PCR product hybridizes to
probe

- PCR primers have biotin group that binds to Streptavidin-horseradish peroxidase
complex

- upon binding to biotin, SA-HRP catalyzes oxidation of 3,3',5,5'-
tetramethylbenzidine (TMB, chromogen) in the presence of H₂O₂ to generate blue
precipitate (chromophores) on membrane = colorimetric quant.

- anywhere that probe binds target, SA-HRP complex binds and yields blue dot on
nylon strip

- Tests for 7 alleles at 1 locus, not very informative on its own

- given that [n(n+1)]/2:

◦ [7(7+1)]/2 = 28 theoretical genotypes for DQα

why is quantification important? - Answer✔️✔️-1) It's a required standard
associated with accreditation by SWGDAM, FBI, ANAB, etc.

2) Protects integrity of DNA extract (a form of evidence)

3) Enhances the possibility that extracted DNA will be available for defense to
perform independent testing

4) Better allows for appropriate amount of DNA so downstream methods are more
successful; very narrow window of [DNA] required for sufficient results

Quantiblot - Answer✔️✔️-hybridization and end point PCR assay; relatively
accurate, expensive



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, ©PREP4EXAMS2024/2025 REAL EXAMS DUMP Wednesday, August 7, 2024 9: 33 PM


- all levels of DNA quality, human specific

1) small sample of extracted DNA is deposited and fixed on nylon membrane
(RLFP path)

2) biotinylated human probe is hybridized to DNA fixed on membrane

3) through colorimetric/chemiluminescent-based process, intensity of developed
image = quantity of DNA in extract

- more DNA = more probe binds = brighter report

- amplifies a target and assumes other samples are equal

Quantifiler - Answer✔️✔️-real-time qPCR assay

- Targets untranslated, intronic hTERT (human telomerase reverse transcriptase)
gene sequence for autosomal DNA and Y-chromosomal DNA for sex
determination

- After each round of PCR, a tungsten-halogen lamp excites free reporter dye and
fluorescence emission captured with CCD camera

- Digitally graphs fluorescent signal over cycle #, allows for estimation of DNA
quantity

- All levels of DNA quality, human-specific; 2x as sensitive as Quantiblot

- considered a TaqMan assay

Quantifiler DNA quantification procedure - Answer✔️✔️-1) make quantification
standards; five 10-fold dilutions of 50 ng stock

2) make master mix (from # of rxns x 8µL primer mix and 10µL PCR reaction
mix), add 18 µL to each applicable well

3) add 2 µL of each standard to respective well



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