ADH2 Intron 3 RsaI Polymorphism

Saturday, May 15, 2004

Description
ADH2 Intron 3 RsaI Polymorphism

Procedure
TEMPLATE: Genomic DNA 100ng/reaction volume of 25 祃

PRIMER 1. A2IN3DW3 5' ATA TTT ATT TTA CCC TAA ACT TAT G 3'
PRIMER 2. A2IN3UP2 5' GAG CTA AAA CAT ACT TTG GAT AG 3'

10 礛 each primer

dNTPs: Final concentration 200礛

BUFFER: Final concentration 2.2mM MgCl2, 50mM KCl, 10mM Tris HCl pH 8.4

TAQ DNA Polymerase: 1unit (Perkin-Elmer Amplitaq DNA polymerase 5u/?l, Cat No. N808-0145)

PCR Protocol: 94oC (5')
94oC (40)", 59oC (30"), 72oC (1.0') ?30 cycles
72oC (10')

Product Size before ------> after RsaI digestion:
RsaI
297 bp --------------> 178 bp and 119 bp

NOTE: Partial intron3 sequence of ADH2 is in GenBank accession #AF040967. The polymorphic nucleotide is 790 bp downstream of exon3 at position 956 of that GenBank entry.

Protocol modified from a RFLP protocol described by Moyra Smith ("Genetics of human alcohol and aldehyde dehydrogenases" Adv. Hum. Genet. 15:249-290, 1986).

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ADH2 Intron 3 RsaI Polymorphism