forensic molecular biology ch 7-9

polymerase chain reaction

polymerase chain reaction

the exponential amplification of specific sequences of DNA to yield sufficient amplified product for downstream applications

0.5 ng

PCR can amplify fragments as small as _______

primer sets

the component of PCR that determines what area of DNA will be copied

forward primer

the primer for PCR that is complementary to the 5' to 3' strand

reverse primer

the primer for PCR that is complementary to the 3' to 5' strand

double stranded

taq polymerase can only attach to ___________ DNA

thermus aquaticus

taq ploymerase is named after...

processivity

the number of bases taq polymerase can add before it falls off

fidelity

the accuracy of taq polymerase

5' to 3'

taq polymerase only has proofreading capability in the ______ direction

hot start PCR

when taq polymerase isn't added until the first annealing step

taqgold

a form of taq polymerase that has been modified and can only be activated once activated at 95ºC

tris buffer

the buffer used that is pH sensitive with temperature variation

multiplex kits

PCR kits that simultaneously amplify 2 or more regions of DNA

94

the denaturation step of PCR is done at _____ºC

60

the annealing step of PCR is done at _____ºC

72

the extension step of PCR is done at _____ºC

95

the pre-incubation step of PCR is done at ____ºC

final extension

the stage of PCR in which taq polymerase adds an extra adenine to each strand

amp+

control sample that ensures the amplification components and thermocycler are working properly

amp-

control sample that ensures purity of amplification components

reagent blank

control sample that ensures the purity of extraction components

stochastic effect

when two alleles in a heterozygous individual are unequally detected, resulting in allele dropout

inhibitors

things that interfere with or prevent the DNA amplification process from occurring properly

inhibitors

heme, melanin, and indigo dyes are examples of _______

BSA

protein from cows that improves the activity of enzymes

staff elimination database

a database in which workers are typed at the same loci as samples to identify the source of contamination

micro-satellite regions

another name for STRs/SSRs

highly variable

STRs are good for human identity purposes because they are highly variable

tetranucleotides

the type of STR repeat unit that is analyzed in forensic applications

stutter

caused by taq polymerase slipping off while extending a region and joining back on in the wrong place

degraded

shorter allele lengths in shorter nucleotide patterns are better for _________ DNA

5' to 3'

STR nomenclature is designated by the number of repeat units on the ______ strand

non-consensus alleles

STR alleles that contain an incomplete repeat unit that fall in between alleles with a full repeat unit. ex.) (GATA)(GAT-)(GATA)

compound repeats

comprise of two or more adjacent simple repeats. ex.) (GATA)(GATA)(GACA)

complex repeats

contain several repeat blocks of variable unit length. ex.) (GATA)(GACA)(CA)(CATA)

THO1

an example of a non-consensus allele, which contains of nine tetranucleotide repeats with one trinucleotide sequence

native electrophoresis

electrophoresis that is done while the DNA is in double stranded form

denaturing electrophoresis

electrophoresis that is done while the DNA is in single stranded form

resolution

denaturing electrophoresis can provide better _________ due to increased flexibility of the single stranded molecule

formamide or urea

in denaturing electrophoresis, ________ is added to denature the DNA. A hydrogen bond is formed with each strand of DNA so the DNA strands do not rejoin. The sample is heated at 95ºC and then the sample is snap cooled.

agarose

electrophoresis gel that has larger pores, separates molecules with sizes of 600-23,000 nucleotides

acrylamide

electrophoresis gel that has smaller pores, separates molecules with sizes of 100-1000 nucleotides

vertically

acrylamide gels are run __________

horizontally

agarose gels are run ___________

capillary

a length of glass filled with polymer that the sample is run through for CE

polymer

what fills the capillary in capillary electrophoresis

performance optimized polymer

POP stands for...

POP4

the polymer used for fragment analysis is...

electrokinetic injection

the sample enters the capillary via __________, where a voltage is applied while the end of the capillary is immersed in a sample. the current pulls negatively charged DNA into the capillary

argon ion laser

the component of the capillary electrophoresis that excites the fluorescent dyes in the sample

internal size standard

a molecular size marker that is added to the sample to permit calibration of sizing measurements

fluorophore

molecule capable of fluorescence

400-600

for DNA typing, dyes fluoresce in the visible range, which is ______ nm

molar extinction coefficient

a fluorophore's ability to absorb light

quantum yield

a fluorophore's efficiency with which it converts absorbed light to emitted light

photostability

a fluorophore's ability to undergo repeated cycles of excitation without being degraded

spectral calibration

accounts for spectral overlap of fluorescent dyes by subtracting out contribution of other dyes for each fluorescent dye, generating a matrix using standards

allelic ladder

a sample containing a mixture of all of the common alleles in a population at specific loci

allelic ladder

converts size of fragments into allele repeat number

mobility modifying non-nt linkers

hexaethylenoxide imparts a shift of ~2.5 nucleotides for each unit, slowing down DNA fragments that may overlap otherwise

amelogenin

an allele that identifies the gender of individual

5 dye system

new identifiler technology that enables smaller PCR products to be generated instead of extending the size of the amplicons