Institute of Plant Genetics, Polish Academy of Sciences, Strzeszyńska 34, 60-479 Poznań, Poland.
BMC Genomics. 2013 Feb 5;14:79. doi: 10.1186/1471-2164-14-79.
The narrow-leafed lupin, Lupinus angustifolius L., is a grain legume species with a relatively compact genome. The species has 2n = 40 chromosomes and its genome size is 960 Mbp/1C. During the last decade, L. angustifolius genomic studies have achieved several milestones, such as molecular-marker development, linkage maps, and bacterial artificial chromosome (BAC) libraries. Here, these resources were integratively used to identify and sequence two gene-rich regions (GRRs) of the genome.
The genome was screened with a probe representing the sequence of a microsatellite fragment length polymorphism (MFLP) marker linked to Phomopsis stem blight resistance. BAC clones selected by hybridization were subjected to restriction fingerprinting and contig assembly, and 232 BAC-ends were sequenced and annotated. BAC fluorescence in situ hybridization (BAC-FISH) identified eight single-locus clones. Based on physical mapping, cytogenetic localization, and BAC-end annotation, five clones were chosen for sequencing. Within the sequences of clones that hybridized in FISH to a single-locus, two large GRRs were identified. The GRRs showed strong and conserved synteny to Glycine max duplicated genome regions, illustrated by both identical gene order and parallel orientation. In contrast, in the clones with dispersed FISH signals, more than one-third of sequences were transposable elements. Sequenced, single-locus clones were used to develop 12 genetic markers, increasing the number of L. angustifolius chromosomes linked to appropriate linkage groups by five pairs.
In general, probes originating from MFLP sequences can assist genome screening and gene discovery. However, such probes are not useful for positional cloning, because they tend to hybridize to numerous loci. GRRs identified in L. angustifolius contained a low number of interspersed repeats and had a high level of synteny to the genome of the model legume G. max. Our results showed that not only was the gene nucleotide sequence conserved between soybean and lupin GRRs, but the order and orientation of particular genes in syntenic blocks was homologous, as well. These findings will be valuable to the forthcoming sequencing of the lupin genome.
窄叶羽扇豆（ Lupinus angustifolius L.）是一种豆科粮食作物，其基因组相对较小。该物种具有 2n=40 条染色体，基因组大小为 960 Mbp/1C。在过去的十年中，窄叶羽扇豆的基因组研究取得了多个里程碑式的进展，例如分子标记的开发、连锁图谱和细菌人工染色体（BAC）文库。在这里，综合利用这些资源来鉴定和测序基因组中的两个富含基因区域（GRR）。
利用与拟茎点霉茎枯病抗性相关的微卫星片段长度多态性（MFLP）标记的序列代表物对基因组进行筛选。通过杂交选择的 BAC 克隆进行了限制性指纹图谱分析和丛集组装，共测序和注释了 232 个 BAC 末端。通过 BAC 荧光原位杂交（BAC-FISH）鉴定了 8 个单基因座克隆。基于物理作图、细胞遗传学定位和 BAC 末端注释，选择了 5 个克隆进行测序。在与 FISH 杂交到单基因座的克隆序列中，鉴定到两个大的 GRR。这些 GRR 与 Glycine max 重复基因组区域具有强烈而保守的同线性，表现在基因顺序和取向完全一致。相反，在具有分散 FISH 信号的克隆中，超过三分之一的序列是转座元件。已测序的单基因座克隆被用来开发 12 个遗传标记，将与适当连锁群相连的窄叶羽扇豆染色体数量增加了 5 对。
一般来说，源自 MFLP 序列的探针可以辅助基因组筛选和基因发现。然而，此类探针对于定位克隆并不有用，因为它们往往会与许多基因座杂交。在窄叶羽扇豆中鉴定的 GRR 中含有较少的分散重复序列，与模式豆科植物 Glycine max 的基因组具有高度的同线性。我们的研究结果表明，不仅大豆和羽扇豆 GRR 中的基因核苷酸序列是保守的，而且在同源性的合成块中特定基因的顺序和取向也是同源的。这些发现将对羽扇豆基因组的即将测序工作具有重要价值。
