Submitted to: Molecular Breeding
Publication Type: Peer reviewed journal
Publication Acceptance Date: 9/1/2005
Publication Date: 11/1/2005
Citation: Hyman, J.R., Simon, P.W. 2005. Sequence Tagged Microsatelite Sites for Carrot (Daucus Carota L.). Molecular Breeding. 16:1-10. Interpretive Summary: Molecular DNA markers allow plant breeders to select plants with the desired genetic material while they are in the seedling stage, reducing the size and expense of field trials as well as the time to market. The majority of molecular DNA markers currently available in carrot indicate whether a fragment of DNA is present but cannot discriminate between the presence of one copy or two of the desired fragment. We have developed molecular DNA markers that can be used to determine whether there are zero, one or two copies of a specific DNA fragment in an individual plant. These markers are of interest to carrot geneticists, breeders and growers who want to decrease the expense and time required to bring new carrot varieties to market.
Technical Abstract: The efficiency of linkage mapping is greatly reduced when dominant molecular markers are used. However, few co-dominant molecular markers are available for use in genetic studies on carrot. We have created plasmid libraries enriched for carrot microsatellites as an initial step in generating sequence tagged microsatellite sites (STMS) for use as co-dominant markers. Single and serial enrichments for di-nucleotide repeats (AC) and (AG) were prepared from DNA cut with five restriction enzymes. Of 331 sequenced clones, 200 contained unique fragments of which 178 contained microsatellites. Although the library was simultaneously enriched for (AC) and (AG), (AG)4+ was much more prevalent (120 clones) than (AC)4+ (75 clones). Regions flanking the (AC) or (AG) repeats contained numerous di-nucleotide and mono-nucleotide repeats. Presence of (A)6+ repeats were positively correlated (r = 24.0%) with (AC) but negatively correlated (r = -15.5%) with (AG) repeats whereas presence of (C)6+ repeats were positively correlated (r = 21.2%) with (AG) but negatively correlated (r = -19.3%) with (AC) repeats. The number of enrichment steps was positively correlated with the length of the microsatellites isolated (r = 31.9%) and the number of duplicated fragments (r = 33.8%) obtained from Sch I derived clones. No significant correlation was observed for these parameters for clones derived from the other restriction enzymes used. Twenty four primer sets designed from a sample of sequences containing 10 or more uninterrupted repeat units produced eight scorable STMS in a segregating F2 carrot population. Although two of these STMS were scored as dominant markers due to the stuttering of bands, all eight markers have been mapped using this population. This is the first report of STMS being generated from a plasmid library which can now be used to create additional STMS for increasing the efficiency of mapping and selection of desirable traits in carrot.