LARGE-SCALE, COST-EFFECTIVE SCREENING OF PCR PRODUCTS IN MARKER-ASSISTED SELECTION APPLICATIONS

被引：71

作者：

GU, WK

WEEDEN, NF

YU, J

WALLACE, DH

机构：

[1] CORNELL UNIV,DEPT HORT SCI,GENEVA,NY 14456

[2] CORNELL UNIV,DEPT PLANT BREEDING,ITHACA,NY 14853

来源：

THEORETICAL AND APPLIED GENETICS | 1995年 / 91卷 / 03期

关键词：

RAPDS; ELECTROPHORESIS; ALLELE-SPECIFIC; ASSOCIATED PRIMERS; COMMON BEAN; PEA;

D O I：

10.1007/BF00222974

中图分类号：

S3 [农学（农艺学）];

学科分类号：

0901 ;

摘要：

A simple, PCR-based method has been developed for the rapid genotyping of large numbers of samples. The method involves a alkaline extraction of DNA from plant tissue using a slight modification of the procedure of Wang et al. (Nucleic Acids Res 21:4153-4154, 1993). Template DNA is amplified using allele-specific associated primers (ASAPs) which, at stringent annealing temperatures, generate only a single DNA fragment and only in those individuals possessing the appropriate allele. This approach eliminates the need to separate amplified DNA fragments by electrophoresis. Instead, samples processing the appropriate allele are identified by direct staining of DNA with ethidium bromide. Total technician time required for extraction, amplification and detection of 96 samples is about 4 h, and this time requirement can be reduced by automation. Excluding labor, cost per sample is less than $0.40. The method is tested using the codominant isozyme marker, alcohol dehydrogenase (Adh-1) gene in pea (Pisum sativum), and applied to the screening of photoperiod genes in common bean (Phaseolus vulgaris L.).

引用

页码：465 / 470

页数：6

共 18 条

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