Degradation and Contamination

Question 1

Crime scenes and preservation problems

Answer 1

exposure to environment
environmental contaminants
tiny amount of cells

Question 2

Environmental exposure on DNA

Answer 2

water and nucleases break down DNA

Question 3

DNA degradation problems

Answer 3

environmental exposure
visualization on gels
hard to amplify large markers

Question 4

Solutions to degradation problems

Answer 4

STRs
mini-STRs (redesign primers closer to STR- may cause null alleles and can’t multiplex)
SNPs
mtDNA

Question 5

PCR inhibitors

Answer 5

heme, melanin, polysaccharides, bile salts, humic compounds, urea, textile dyes

Question 6

Blood as source

Answer 6

heme

Question 7

Tissue and hair as source

Answer 7

melanin

Question 8

Feces as source

Answer 8

polysaccharides and bile salts

Question 9

urine as source

Answer 9

urea

Question 10

clothing as source

Answer 10

textile dyes

Question 11

PCR inhibitors causes

Answer 11

interference at multiple steps from cell lysis to polymerase activity
lead to preferential amplification

Question 12

Solutions for inhibitors

Answer 12

dilute DNA template
add more Taq (or different polymerase)
add additive
include a separation step

Question 13

Ways to increase strength of DNA signal

Answer 13

increase number of PCR cycles
reduce PCR volume
increase amount of DNA
adjust chemistry in capillary electrophoresis
increase injection time for capillary electrophoresis

Question 14

Issues with low copy number

Answer 14

allele drop in
allele drop out
greater stutter
peak imbalance

Question 15

Solutions for low copy number

Answer 15

PCR every sample 2x

sterile labs

Question 16

Contamination sources

Answer 16

environment (evidence collection team, manufacturers)
other samples/technicians
other samples after PCR

Question 17

Solutions to prevent contamination

Answer 17

standard operating protocols
designated work areas
negative controls
staff elimination database
sterile labs

Question 18

Sterile labs

Answer 18

disposable gloves, masks coats

all surfaces bleached regularly