Park J Y, Koo D H, Hong C P, Lee S J, Jeon J W, Lee S H, Yun P Y, Park B S, Kim H R, Bang J W, Plaha P, Bancroft I, Lim Y P
Department of Horticulture, Chungnam National University, Kung-Dong 220, Yusong-Gu, Daejeon 305-764, South Korea.
Mol Genet Genomics. 2005 Dec;274(6):579-88. doi: 10.1007/s00438-005-0041-4. Epub 2005 Nov 9.
We constructed a bacterial artificial chromosome (BAC) library, designated as KBrH, from high molecular weight genomic DNA of Brassica rapa ssp. pekinensis (Chinese cabbage). This library, which was constructed using HindIII-cleaved genomic DNA, consists of 56,592 clones with average insert size of 115 kbp. Using a partially duplicated DNA sequence of Arabidopsis, represented by 19 and 9 predicted genes on chromosome 4 and 5, respectively, and BAC clones from the KBrH library, we studied conservation and microsynteny corresponding to the Arabidopsis regions in B. rapa ssp. pekinensis. The BAC contigs assembled according to the Arabidopsis homoeologues revealed triplication and rearrangements in the Chinese cabbage. In general, collinearity of genes in the paralogous segments was maintained, but gene contents were highly variable with interstitial losses. We also used representative BAC clones, from the assembled contigs, as probes and hybridized them on mitotic (metaphase) and/or meiotic (leptotene/pachytene/metaphase I) chromosomes of Chinese cabbage using bicolor fluorescence in situ hybridization. The hybridization pattern physically identified the paralogous segments of the Arabidopsis homoeologues on B. rapa ssp. pekinensis chromosomes. The homoeologous segments corresponding to chromosome 4 of Arabidopsis were located on chromosomes 2, 8 and 7, whereas those of chromosome 5 were present on chromosomes 6, 1 and 4 of B. rapa ssp. pekinensis.
我们用大白菜(Brassica rapa ssp. pekinensis)的高分子量基因组DNA构建了一个细菌人工染色体(BAC)文库,命名为KBrH。该文库使用经HindIII酶切的基因组DNA构建,由56,592个克隆组成,平均插入片段大小为115 kbp。我们利用拟南芥中一个部分重复的DNA序列(分别由第4和第5号染色体上的19个和9个预测基因代表)以及KBrH文库中的BAC克隆,研究了大白菜中与拟南芥区域相对应的保守性和微同源性。根据拟南芥同源基因组装的BAC重叠群揭示了大白菜中的三倍化和重排。一般来说,旁系同源片段中基因的共线性得以维持,但基因含量因间隙缺失而高度可变。我们还使用组装重叠群中的代表性BAC克隆作为探针,通过双色荧光原位杂交技术,将它们与大白菜的有丝分裂(中期)和/或减数分裂(细线期/粗线期/减数第一次分裂中期)染色体进行杂交。杂交模式从物理上鉴定了大白菜染色体上拟南芥同源基因的旁系同源片段。与拟南芥第4号染色体相对应的同源片段位于大白菜的第2、8和7号染色体上,而与第5号染色体相对应的同源片段则存在于大白菜的第6、1和4号染色体上。