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United States Department of Agriculture

Agricultural Research Service

Title: A Novel DNA Assay for Marker-Assisted Breeding

Authors
item Reddy, A - TEXAS A&M UNIV
item Ayres, Nicola - TEXAS A&M UNIV
item Larkin, Patrick - TEXAS A&M UNIV
item Park, William - TEXAS A&M UNIV
item McClung, Anna
item McClung, Anna
item Pfeifer, J - TEXAS A&M UNIV

Submitted to: Rice Technical Working Group Meeting Proceedings
Publication Type: Proceedings
Publication Acceptance Date: February 1, 1996
Publication Date: N/A

Technical Abstract: A number of important genes have already been tagged in crop plants and work on many more is currently underway. However, the application of DNA diagnostics in plant breeding is still in its infancy. Much of this work has been done using RFLP and other technologies cumbersome to apply to the thousands of samples that a breeder typically examines in a season. Molecular breeding programs thus require DNA assays that are rapid, cost effective and can be performed on thousands of samples. We are attempting to develop a cost effective high through-put DNA marker technology which does not require gel electrophoresis to facilitate marker-assisted breeding in plants. The TaqMan assay was tested using the granule bound starch synthase (GBSS) gene of rice, which is largely responsible for amylose content. Amylose content is important in rice as it largely determines rice cooking quality. Currently, about 20,000 amylose assays are performed each year on breeding lines developed by US public rice breeders. We have confirmed a single nucleotide change adjacent to the putative 5' leader intron splice site of GBSS gene by sequencing 32 rice varieties. The sequence data demonstrated that all 17 intermediate and high amylose varieties had the sequence ACGT in the putative 5' leader intron splice site while all 15 low amylose varieties had the sequence ACTT. TaqMan probes were produced corresponding to both alleles. PCR was performed on these 32 varieties, and also a segregating F5 population derived from a cross between low and high amylose varieties. Results obtained using this new non-gel based diagnostic technique indicated that it was effective in identifying the GBSS gene in rice and suggests that this will be an important tool for use in marker-assisted plant breeding.

Last Modified: 12/19/2014