Lab 8-10 | PCR, Agarose Gel Electrophoresis

Chelex

A combination of chelating agents that bind to cations in solution. The purpose of this is to protect DNA from degradation by binding to metal ions that are essential for nuclease activity

Polymerase Chain Reaction (PCR)

Amplifying small amounts of DNA into larger amounts

Template

gDNA; harvested from cheek tissue (Buccal cells)

Primers

Approximately 20 nucleotides of DNA for the PCR to begin 

dNTPs

Deoxynucleotide triphosphate; essential building blocks of DNA

Taq polymerase

Synthesizes new DNA strands by extending a primer in the 5’ → 3’ direction

1. Denaturing

Melting double stranded DNA at 95°C into single stranded DNA

2. Annealing

Binding primers to DNA using molecular complementarity at 55°C

3. Extension

Replication occurs at 72°C

TAS2R38

A taste receptor that can taste bitter phenylthiocarbamide (PTC)

T¹ → Taster

T² → Nontaster

T¹T² → Intermediate taster

Buffers

Mimics cytoplasmic conditions

Consensus sequence

A specific nucleotide sequence that is recognized by an enzyme

Restriction enzyme

Digest DNA

Hae III

An endonuclease that recognizes the sequence in SNP between taste and nontaster by digesting taster alleles

Bioinformatics

Analyzing biological data using computer algorithms

Cite Directed Mutagenesis (CDM)

A laboratory method used to create specific, targeted changes, such as insertions, deletions, or substitutions

SNP in TAS2R38

C to G alteration