Kidd Lab 

HindIII (B) site (located in original G8 clone, about 0.5 kb downstream (centromeric) of pKO-82)

Template: Genomic DNA 100-200 ng/reaction volume of 20 µl

Primer 1.     PKOH3BF:     5'-GCA CCT GGA TCT CGG GCT TC-3'        100 ng/reaction
Primer 2.     PKOH3BR:     5'-GCC GGC CAG ATT GGT GTC C-3'         100 ng/reaction

dNTPs: Final Concentration 200 µM/reaction

Buffer: Final Concentration 1.5 mM MgCl₂, 50 mM KCl, 10 mM Tris-HCl pH 8.4

Taq DNA Polymerase: 0.5 Units/reaction
                                 (Perkin-Elmer Amplitaq DNA polymerase 5 U/µl, Cat. No. N808-0145)

PCR Cycling Profile: 95^oC (5')
                              95^oC (30"), 62^oC (30"), 72^oC (30")  × 30 cycles

Product size:
                                HindIII
                   194 bp---------------> 118 and 72 bp (for homozygote site present)

Reference: Stapleton, P.A., 1988. Nucleic Acids Res. 16:2735.
 
 

Kidd Lab Histograms for Allele Frequencies