SEQUENCE-TAGGED-SITE-FACILITATED PCR FOR BARLEY GENOME MAPPING

被引:108
作者
TRAGOONRUNG, S
KANAZIN, V
HAYES, PM
BLAKE, TK
机构
[1] MONTANA STATE UNIV,DEPT PLANT & SOIL SCI,BOZEMAN,MT 59717
[2] OREGON STATE UNIV,DEPT CROP SCI,CORVALLIS,OR 97331
关键词
SEQUENCE-TAGGED-SITE; POLYMERASE CHAIN REACTION; POLYACRYLAMIDE-GEL ELECTROPHORESIS; 4-BASE CUTTER;
D O I
10.1007/BF00227417
中图分类号
S3 [农学(农艺学)];
学科分类号
0901 ;
摘要
Speed, efficiency, and safety considerations have led many genome mapping projects to evaluate polymerase chain reaction (PCR) sequence amplification as an alternative to Southern blot analysis. However, the availability of informative primer sequences can be a limiting factor in PCR-based mapping. An alternative to random amplified polymorphism detection (RAPD) is the sequence-tagged-site (STS) approach. If informative primer sequences could be derived from known sequences, then current maps, which are based on both known function and anonymous clones, might be easily converted to maps utilizing PCR technology. In this paper, four pairs of primer sequences were obtained from published sequences, and four pairs were obtained by sequencing portions of DNA clones from genomic clones derived from a random genomic library used in the North American Barley Genome Mapping Project (NABGMP). These primers were used to screen for polymorphisms in the progeny of a winter x spring and a spring x spring barley cross. Two types of polymorphisms were distinguished using these primer sets: (1) insertion/deletion events that could be read directly from agarose gels, and (2) point mutation events. The latter were identified using polyacrylamide-gel electrophoresis of PCR products following digestion with restriction endonucleases (four-base cutters). To determine whether the PCR-based. polymorphisms were allelic to polymorphisms identified by the clones from which the primer sequences derived, chromosomal assignments and (when possible) co-segregation analysis was performed.
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页码:1002 / 1008
页数:7
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