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United States Department of Agriculture

Agricultural Research Service

Title: Rapid Development of Gene-Tagged Microsatellite Markers from Bac Clones of Channel Catfish, Ictalurus Punctatus, and Rainbow Trout, Oncorhynchus Mykiss

item Waldbieser, Geoffrey
item Karsi, Attila - USDA, ARS, CGRU
item Gahr, Scott
item Rexroad, Caird

Submitted to: Genetics in Aquaculture Symposium
Publication Type: Abstract Only
Publication Acceptance Date: November 9, 2003
Publication Date: November 9, 2003
Citation: Waldbieser, G.C., Karsi, A., Gahr, S.A., Rexroad III, C.E. 2003. Rapid development of gene-tagged microsatellite markers from bac clones of channel catfish, ictalurus punctatus, and rainbow trout, oncorhynchus mykiss. Genetics in Aquaculture Symp. VII. Nov 9-15, 2003. p.22.

Technical Abstract: Polymorphic markers linked to conserved genes (type I markers) permit identification of alleles linked to candidate genes, placement of conserved genes on genetic linkage maps, and integration of linkage and physical maps. A technique was developed to improve the efficiency of producing gene-linked microsatellite markers in channel catfish and rainbow trout. Template DNA was prepared from cultures derived from single BAC colonies using a simple alkaline lysis miniprep. Presence of conserved genes in each BAC clone was verified by sequencing with gene-specific primers. The BAC templates were directly sequenced using primers consisting of ten tandem repeats (TAA repeats for catfish, TC or TG repeats for trout) with one or two unique 3' terminal bases. These primers were termed STRAPs for 'Short Tandem Repeat-Anchored Primers'. At least one STRAP provided sufficient 3' flanking sequence from each clone for design of a BAC-specific primer. The BAC-specific primer was used to sequence back through the tandem repeat and obtain 5' flanking sequence, and a second BAC-specific primer was designed for microsatellite genotype analysis. The catfish markers contained an average of 15 trinucleotide repeats, and trout markers contained an average of 18 dinucleotide repeats. All markers demonstrated allelic polymorphism in reference populations. This technique provided a rapid method for production of type I markers in two fish genomes.

Last Modified: 3/1/2015
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