Lab 3.4 2012

M 3/26/12

CORRECT THE VOLUMES IN THE LAB MANUAL 21 mL 10X in 210 mL total!

Goal for this lab: Genotype Salk line plants, looking for homozygous mutants.

Use 3 primers in a single reaction (GSP1, GSP2, LBP)

LB 195 LBb1.3 ATTTTGCCGATTTCGGAAC (111 bp from left border)

Materials

• PCR strip tubes
• Harris cutting mat
• Harris Uni-Core (0.5 mm) Cutting Tool
• Beakers for bleach

Reagents

• 2% bleach (12 mL bleach + 588 mL dH2O)
• 5U/µL Taq (3.5 µL per pair)
• 10X polymerase buffer (21 + 21 = 42 µL per pair)
• 2.5 mM dNTP mix (51 µL per pair)
• Sterile dH2O
• Working stocks of primers, including LBP diluted to 10 µM (17µL/rxn)