Submitted to: Molecular Genetics and Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/23/2003
Publication Date: 10/1/2003
Citation: LI, L., LU, S., O'HALLORAN, D.M., GARVIN, D.F., VREBALOV, J. HIGH-RESOLUTION GENETIC AND PHYSICAL MAPPING OF THE CAULIFLOWER HIGH BETA-CAROTENE OR (ORANGE) GENE. MOLECULAR GENETICS AND GENOMICS. 2003. v. 270. p. 132-138. Interpretive Summary: Vitamin A deficiency is a global problem. It is estimated that more than 250 million children worldwide are at risk. As humans are unable to synthesize vitamin A de novo from endogenous isoprenoid precursors, dietary carotenoids are the most potent precursor of vitamin A. To enhance the nutritional value of plant foods via increased carotenoid content, we need to understand how carotenoid synthesis and accumulation are regulated in plants. A cauliflower Or gene mutant which induces the accumulation of high levels of B-carotene provides a good model to study regulation of these processes. In this paper, as part of a map-based cloning strategy,we describe high-resolution genetic mapping of the Or gene, the construction of a cauliflower large-insert DNA library, and the establishment of the region of the genome containing this gene. Based on this research, we were able to narrow the location of the Or gene to a very small DNA fragment. The results reported here provide the essential information to isolate this interesting and potentially important Or gene in improving carotenoid content in crops.
Technical Abstract: The cauliflower Or gene causes high levels of beta-carotene to accumulate in various tissues of the plant that are normally devoid of carotenoids. To decipher the molecular basis by which Or regulates carotenoid accumulation, we have undertaken the isolation of Or by a map-based cloning strategy. Two previously isolated locus-specific sequence-characterized amplified region (SCAR) markers that flank Or were employed for the analysis of a large segregating population consisting of 1632 F2 individuals, and a high-resolution genetic linkage map of the Or locus region was developed. To facilitate positional cloning, we constructed a cauliflower bacterial artificial chromosome (BAC) library using high molecular weight DNA from the Or homozygous plants. The BAC library comprises 60,288 clones with an average insert size of 110 kb and represents an estimated 10-fold coverage of the genome. A BAC contig encompassing the Or locus was established by screening the library with a marker closely linked to Or and by overlapping BAC clones through chromosome walking. Physical mapping delimited the Or locus to a 50-kb DNA fragment within a single BAC clone, which corresponds to a genetic interval of 0.3 cM.