Technical Oral Exam Flashcards

Question

describe the electrophoretic flow vs. the electroosmotic flow

Answer 1

EPF - DNA (-) moves away from the cathode (-) towards the anode (+) EOF - opposite of the EPF, the anode (+) buffer flows towards the cathode (-)

Answer 2

protein added to help stabilize polymerase reduces/prevents PCR inhibition

Answer 3

1. DNA (template DNA) 2. primers 3. deoxynucleoside triphosphates (dNTPs) 4. DNA polymerase 5. bovine serum albumin (BSA) 6. magnesium chloride 7. potassium chloride 8. tris-HCl buffer

Answer 4

autosomal DNA - 84 bp Y (male) DNA - 81 bp & 136 bp degradation - 294 bp inhibition - 435 bp

Answer 5

CXR dye present in all wells of rxn plate at same concentration purpose - normalization; differentiate background noise of fluctuations of instrument from the fluorescence levels of DNA in sample

Answer 6

graph of the Cq values of the standards vs. the Log of the concentration of the standards slope (regression line) indicates PCR efficiency regression line is calculated using best fit curve with quant standard data points utilizes standard dilution series used to determine the concentration of unknown DNA samples (software calculates DNA concentration based on Ct/q value of sample & comparing it to the standard curves Ct/q's

Answer 7

Ct/q = m(logQty)+b Qty = starting amount of DNA

Answer 8

4 1. 50 ng/ul 2. 2 ng/ul 3. 0.08 ng/ul 4. 0.0032 ng/ul

Answer 9

the expected Ct/q value for a sample with a quantity of 1 ng

Answer 10

measure of closeness of fit between the standard curve regression line & individual Ct/q data points of standards 1 perfect fit / >/= 0.99 passing R2 value

Answer 11

PCR efficiency -3.3 100% efficient / -3.1 - -3.6 passing slope

Answer 12

made from purified whole blood from multiple male donors expires after 7 days critical for accurate analysis of run data stock concentration is 50ng/ul

Answer 13

1. spatial resolution 2. spectral resolution 3. DNA sizing precision

Answer 14

it determines the relationship between the signal emitted from each capillary & where it's detected on the CCD camera - it maps the position of each capillary to a region on the camera necessary to separate DNA fragments that may differ in size by a single nucleotide

Answer 15

helps separate out spectral dye overlap necessary to separate fluorescent dye colors from one another so that dye labeled PCR products from different loci can be resolved

Answer 16

must be consistent from run to run enough so that samples can be related to allelic ladders & yield reproducible results

Answer 17

International Organization of Standards ISO17025 - required for testing & calibration laboratories

Answer 18

Amended Reports AR3125 - required for forensic testing & calibration laboratories

Answer 19

Quality Assurance Standards FBI QAS - set forth by FBI for DNA testing laboratories

Answer 20

2 one for forensics and one from databasing

Answer 21

17 main standards

Answer 22

Scientific Working Group on DNA Analysis Methods publishes guidelines or recommendations for best practices in forensic DNA laboratories - how to best implement ISO & QAS standards into your lab

Answer 23

isolation of two cell types - epithelial cells & sperm cells to extract DNA accomplished by selectively lysing non-sperm cells & removing the non-sperm DNA prior to lysing sperm cells sperm have stronger cell membranes than epithelial cells & in turn are more difficult to lyse

Answer 24

1. there may be more epithelial cells than sperm cells 2. incomplete initial digestion 3. inadequate washing of sperm 4. carryover of F1 into F2

Answer 25

dithiothreitol reduces disulfide bonds of proteins in cell membranes allowing release of the DNA especially crucial for digesting hair and sperm cells

Answer 26

proteinase K serine protease - cleaves peptide bonds in proteins imbedded in cell membranes enzyme used to digest proteins (histones) to expose the DNA inactivates nucleases that break down DNA

Answer 27

tissue lysis buffer lysis of cell membranes to release & expose DNA

Answer 28

carrier RNA enhances binding of DNA to surface of magnetic particles, especially hen low amounts of DNA is expected drives binding process added to increase DNA yield in low concentration samples

Answer 29

tissue lysis buffer lysis of cell membranes to release & expose DNA less harsh than ATL! used in differential extractions to selectively lyse epithelial cells before sperm cells

Answer 30

lysis buffer with chaotropic salts used together with EZ2 for automated purification large volume samples

Answer 31

GMID-X utilizes our ILS (WEN) to size unknown DNA fragments it takes 4 fragments closest to the unknown fragment & creates two size curves the first size curve utilizes two peaks before & one peak after the second size curve utilizes one peak before and two peaks after it takes the average of these two size curves & gives you the base pair size of our unknown DNA fragment

Answer 32

x-axis - plots the base pair size of DNA fragments y-axis - plots the rfu value (concentration of DNA present)

Answer 33

GMID-X utilizes the raw data from our CE runs to plot it onto an electropherogram so we can easily read it. it plots the size of the DNA fragments (by number of base pairs; x-axis) and the concentration of the DNA (rfu value; y-axis)

Answer 34

an allele present in the sample, yet it is not amplified often caused by primer binding sit mutations (if the primer doesn't bind, amplification does not occur)

Answer 35

defines the height requirement at and above which detected peaks can be reliably distinguished from background noise (100rfu)

Answer 36

only used when pull-up is seen at 100rfu upper rfu threshold which should only be applied to a sample exhibiting high peak heights where baseline noise or pull-up is present above the AT causing artifacts to be labeled as true peaks (300rfu)

Answer 37

peak height or signal magnitude below which it is reasonable to assume that allelic DO of a sister allele in a heterozygous pair may have occurred (350rfu) ST was determined during validation studies using sensitivity data (known profiles at decreasing concentrations)

Answer 38

1. determine the minimum number of contributors 2. maj/min or indistinguishable? 3. interpretable or uninterpretable? 4. designate genotypes 5. utilize available assumptions for further resolution of genotypes if appropriate 6. compare evidence to applicable references 7. perform statistical evaluation if applicable

Answer 39

p2 + 2pq + q2 = 1 mathematical relationship between allele frequencies and genotype frequencies principle of a perfectly balanced population where the genetic variation remains constant between generations in the absence of disturbing factors HWE is dependent on Mendel's law of segregation & independent assortment of genes during sex cell formation

Answer 40

provides a stable pH environment (pH can alter fidelity of a dNTP insertion) pH between 8.3-8.8 is used (@ 20C) - pH decreases to 6.9 when heated to 96C to activate Taq

Answer 41

refers to a stable, independent, and random flow of alleles within a population directly related to the ability to achieve or approach HWE & uses the product rule to combine locus probabilities LE is also dependent on Mendel's law of segregation & independent assortment of genes during sex cell formation

Answer 42

correction factor used to address the issue of increased homozygosity from a population substructure accounts for the difference between population genotype frequencies & true HWE 0.1 or 0.3

Answer 43

homo - could be unjustly prejudicial towards a defendant, especially in a smaller population, by raising a statistic hetero - could benefit a defendant by lowering a statistic

Answer 44

setting theta too high = overcorrect & possibly reduce the statistic inappropriately (too much) setting theta too low = undercorrect & possibly reduce the statistic insufficiently (too little)

Answer 45

overall, the rarity of the genotype is REDUCED with increasing theta values in homozygous loci (more conservative). for heterozygous loci, the rarity is INCREASED with increasing theta values (less conservative) thus, theta is only applied to homozygous loci & not heterozygous loci in order to calculate a more conservative estimate

Answer 46

heterozygous: f = 2pq homozygous: f = p2 +p(1-p)*theta value* - instead of p2

Answer 47

statistical principle allowing unlinked or independent events to be combined using multiplication for unrelated events (independent), such as locus probabilities of a DNA profile, the probabilities for each locus can be combined using the product rule ("and" means multiplication) if two events are independent (A & B have no effect on each other), we can calculate the probability of A AND B by multiplication

Answer 48

minimum allowable frequency within a population group & is based on the size of the population sample used for unobserved alleles & raising the frequencies that fall below the MAF if the allele is observed fewer than 5 times, Popstats will assign a MAF of 5/2n, which assumes a minimum of 5 observations of that allele in the sample size 'n'

Answer 49

5 / 2n the numerator (5) is the minimum number of times that an allele should be seen for a reliable frequency the denominator (2n) is 2 multiplied by size of the database to account for all observed alleles within a locus

Answer 50

if an allele is rare (observed few or 0 times in the sample database), we must provide a conservative estimate for use in statistics - so we don't skew the statistic too much ( we need to be conservative)

Answer 51

this means looking at the mixture to attempt to narrow down the possibilities for the subject peak heights are utilized to account for all reasonable genotypes that would explain the unknown Popstats is not sophisticated enough to do this

Answer 52

this means that all possibilities are considered of the deduced profile regardless of the characteristics of the mixture using the unrestricted LR would account for all possible genotypes that could explain the unknown this is what Popstats calculates

Answer 53

suppose you have an RMP of 1 in 1 million. the defense may argue, "Louisiana has a population of almost 5 million people. this means there are 4 other people i La alone that could have left that profile"

Answer 54

it requires the assumption that all contributors are fully represented at the loci being utilized (no possibility of DO)

Answer 55

the defendant's fallacy can be manifested from the prosecutor as well: suppose you have an RMP of 1 in 1 quadrillion. the prosecutor may ask, "the world population is approximately 7 billion people. how many worlds would you need to see this profile again?"

Answer 56

the RMP refers to an outcome probability over one trial, not how many times that outcome is expected over several trials so, given that the profile has been observed once already, it describes the probability of observing this profile again if a random person is selected from the population. each time a person is selected, the probability is 1 in *value*

Answer 57

1. random mating 2. large population 3. no gene flow 4. no mutations 5. no natural selection

Answer 58

prevents inbreeding &/or the occurrence of a population substructure (amish people)

Answer 59

ensures allele frequencies are not changed through genetic drift

Answer 60

this would increase variability in the gene pool

Answer 61

to avoid introducing new alleles into a population

Answer 62

stronger genes may be 'chosen' over inferior genes which would cause allele frequencies to change if alleles were being favored or lost

Answer 63

GD - small population (vertical, inbreeding) GF - large population to large population (horizontal)

Answer 64

scientists performed an Exact Test (simulation that compared a population in perfect HWE to our population that's in HWD) & found: 1. no significant deviation from HWE 2. loci are sufficiently discriminatory 3. there are significant differences in allele frequencies among different population groups 4. loci are sufficiently independent (no evidence of association), so the product rule is valid

Answer 65

genetic markers are chosen that are less affected by disruption alleles chosen are from loci for noncoding regions of DNA (not expressed as phenotypes, reducing genetic divergence, & loci chosen are typically spaced far apart or on different chromosome (ensuring independence))

Answer 66

Combined DNA Index System FBI's program of support for criminal justice DNA databases as well as the software used to run these databases purpose: to generate investigative leads for unsolved cases, to identify UHR, & help solve missing persons cases 3 hierarchal levels: NDIS (national), SDIS (state), LDIS (local)

Answer 67

1. forensic unknown 2. forensic partial 3. forensic mixture 4. forensic targeted 5. SDIS forensic 6. SDIS mixture

Answer 68

complete, single source or deduced single source profile

Answer 69

single source or deduced single source incomplete profile MME value of >1E+007

Answer 70

mixture with >3 alleles at two or more loci & has an MME value of >1E+007

Answer 71

MME value of <1E+007 MRE value of >1E+007

Answer 72

single source MME value of >1E+005 MRE value of <1E+007

Answer 73

mixture MME value of >1E+005 MRE value of <1E+007 ^ *same as SDIS forensic*

Answer 74

moderate match estimation calculates estimated match probability value for all specimens in a selected specimen category; not configurable calculated evaluating 13 CODIS core loci at moderate stringency

Answer 75

match rarity estimation calculates estimated rarity value for all specimens in a selected specimen category, has many configurations locus stringency can be changed can be calculated for the 20 expanded loci

Answer 76

match rarity estimation ; stringency MME is match rarity estimate when the 13 original core loci are used & all stringencies are set to moderate MRE is match rarity estimate when you calculate stringency by locus; hetero loci are searched at HIGH stringency; homo loci are searched at MODERATE stringency

Answer 77

during gamete formation, the two alleles at a gene locus segregate from each other; each gamete has an equal probability of containing either allele during meiosis when sex cells divide

Answer 78

DNA Index of Special Concern virtual index of forensic unknowns in CODIS that meet certain eligibility requirements created to facilitate searching of Arrestee Rapid DNA profiles enrolled in CODIS from booking stations

Answer 79

1. homicide, SA, terrorism, kidnapping 2. forensic unknown 3. must be complete at 13 original CODIS core loci 4. source ID = NO 5. must have required casework metadata (investigating agency info, offense description, etc.)

Answer 80

used as a filter to report only locus matches of equal or higher quality *locus stringency - locus to locus* high - exact match moderate - 'fits in the bucket' low - one allele matches, one doesn't *match stringency - profile as a whole* high - all loci match moderate - at least one locus is at moderate; none at low low - at least one locus is at low

Answer 81

when primers interact with each other & bind to each other polymerase can extend the annealed primers to produce primer-dimers which compete for reagents in a PCR reaction

Answer 82

1. lag phase - baseline; before significant product formation 2. exponential phase - close to 100% efficiency, amplicon formation is doubling each cycle 3. linear phase - rxn efficiency slows to an arithmetic increase; PCR components fall below critical concentration 4. plateau phase - product formation has diminished; PCR components have been exhausted

Answer 83

lyse - break open the cells to expose the DNA bind - chaotropic salts introduced via reagents reduce the pH and causes the DNA to bind to the silica beads wash - a system of washes get rid of all of the other components and debris other than the DNA elute - the chaotropic salts are washed away and the pH is raised again causing the DNA to elute into the elution tube