Submitted to: Journal of Theoretical and Applied Genetics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: February 18, 2001
Publication Date: N/A
Interpretive Summary: In breeding for increased beta-carotene content in tomato, traditional methods utilize crosses between cultivated red-pigmented, low beta-carotene plants and orange-pigmented high beta-carotene containing wild species. Using molecular markers linked to a trait of interest, selection can be performed at early seedling stages, and true breeding lines identified with relative ease. We have identified molecular markers linked to the gene Beta for beta-carotene content. These markers are termed RAPD markers. These are simple, fast and inexpensive to use; however, they are sensitive to slight changes in reaction conditions, resulting in poor reproducibility. To overcome this problem, RAPD markers can be converted into a different class of markers termed SCAR markers. SCAR markers are characterized by high specificity for a single gene and their reproducibility is excellent. In this study, we cloned and sequenced two RAPD markers linked to Beta and utilized this information to develop codominant and dominant SCAR markers linked to the Beta gene. The markers can be used by agriculturalists and researchers to facilitate quick and accurate identification and characterization of the Beta gene in tomato and facilitate development of new cultivars with increased beta-carotene content.
Two previously identified RAPD markers, OPAR1067 and UBC792, linked to the Beta (B) locus in tomato were cloned and sequenced. The end sequences were used to design longer allele specific SCAR primer pairs, SCAR18f/r and SCBC92f/r. Each of these amplified a single same sized product as their respective progenitor RAPD markers, but neither differentiated the two parental genotypes under a variety of PCR conditions. The amplified SCAR products from the null phenotype parent were cloned and full-length sequences were obtained. Site mutation searching identified a RsaI site mutation on the allele of the high beta-carotene parent in the SCAR18f/r amplified DNA product. A HinfI site mutation was identified on the allele of the orange-fruited parent in SCBC792f/r amplified DNA product from which a codominant SCBC792779 marker was generated.The SCBC792fl/r amplified a 682-bp fragment that was only present in the high beta-carotene parent and a dominant SCBC792682 marker was generated. Using 50 introgressed lines, all three SCAR markers were mapped to the long arm of chromosome 6, consistent with the location of B on the classical linkage map of tomato.