Map-based cloning of a recessive genic male sterility locus in Brassica napus L. and development of its functional marker

Abstract

We previously mapped one male-sterile gene (Bnms3) from an extensively used recessive genic male sterility line (9012AB) in Brassica napus to a 0.14-cM genomic region. In this study, two highly homologous BAC contigs possibly containing the candidate BnMs3 gene were identified using a map-based cloning strategy. A BnMs3-linked SCAR marker (DM1) capable of differentiating the subgenomes between B. rapa and the B. oleracea aided mapping of BnMs3 on the contig derived from the B. napus chromosome C9. One representative BAC clone was sequenced from each of the two contigs and resulted in a larger number of markers according to the sequence difference between the two clones. To isolate BnMs3, these markers were then analyzed in another two BC(1) populations with different genetic backgrounds. This assay allowed for a delimitation of the mutated functional region of BnMs3 to a 9.3-kb DNA fragment. Gene prediction suggested that one complete open reading frame (ORF, ORF2) and partial CDS fragments of ORF1 and ORF3 reside in this fragment. Sequence comparison and genetic transformation eventually indicated that ORF1 (designated as BnaC9.Tic40), an analogue of the Arabidopsis gene AT5G16620 which encodes a translocon of the inner envelope of chloroplasts 40 (Tic40), is the only candidate gene of BnMs3. Furthermore, two distinct mutation types in ORF1 both causing the male-sterile phenotype were individually revealed from 9012A and the temporary maintainer line T45. The molecular mechanism of this male sterility as well as the application of BnMs3-associated functional and cosegregated markers in true breeding programs was also discussed.

A triallelic genetic male sterility locus in Brassica napus: an integrative strategy for its physical mapping and possible local chromosome evolution around it

“…To differentiate this male-sterile locus from those of other spontaneous GMS lines in B. napus, such as BnMs1 and BnMs2 in S45AB (Pan et al, 1988), BnMs3 and BnMs4 in 9012AB (Chen et al, 1998; possibly identical to the MSL system and the Syngenta-patented NMS system discussed by Li et al, 2012), this locus has been uniformly addressed as BnMs5. Accordingly, the three alleles of this locus are individually assigned as BnMs5 a (restorer type, corresponding to the previous Rf or Mf ), BnMs5 b (male-sterile type, corresponding to the previous Ms) and BnMs5 c (normal male-fertile type, corresponding to the previous ms), with the dominance relationship of BnMs5 a .…”

“…Although the amphidiploid B. napus represents a complicated genome derived from the natural hybridization between ancestors of the diploid B. rapa (A genome donor) and B. oleracea (C genome donor), map-based cloning is still an effective way for the isolation of genes responsible for obvious phenotypic variation v (Yi et al, 2010;Dun et al, 2011;Li et al, 2012). The first step in map-based cloning is the construction of a high-resolution genetic map around the target gene.…”

Screening ofChinese cabbage mutants produced by60Co γ‐ray mutagenesis of isolated microspore cultures

“…Male sterility is an important tool for hybrid seed production, and the exploration of male sterility mechanisms remains a highly active research area (Araya et al 1998, Hanson andBentolila 2004, Cao et al ). In the past decade, numerous genes that control male fertility in plants have been cloned and isolated through the analysis of mutants (Li et al 2012, Shu et al 2012, Igarashi et al 2013, and these studies have helped reveal the mechanisms that regulate plant fertility. However, some mechanisms remain to be elucidated.…”

Common bean reaction to angular leaf spot comprises transcriptional modulation of genes in the ALS10.1 QTL

“…for example, reverse genetic has been shown to be an important tool for gene discovery and functional analysis. However, for species with little or inexistent genomic resources, forward genetics by positional cloning is an efficient alternative technology for gene discovery, especially those controlling traits of agronomic importance in plants (Li et al, 2012; Xia et al, 2012). Phaseolus vulgaris L. (common bean) is one of the latter, and despite its social and economic importance (Broughton et al, 2003; Gepts et al, 2008), common bean genomics has lagged behind other crops.…”