Construction of a whole-genome radiation hybrid panel for high-resolution gene mapping in pigs
Construction of a whole-genome radiation hybrid panel for high-resolution gene mapping in pigs
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DOI：
10.1159/000015095
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发表时间：
1998-01-01
期刊：
影响因子：
--
通讯作者：
Gellin, J
中科院分区：
文献类型：
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作者：
Yerle, M;Pinton, P;Gellin, J
我们已经开发了一个面板的152全基因组辐射杂交融合辐射二倍体猪淋巴细胞或成纤维细胞与受体仓鼠永久细胞。通过用SINE探针进行荧光原位杂交或用SINE特异性引物进行引物原位标记（PRINS）来检查保留在每个杂交克隆中的猪染色体片段的数量和大小。一种策略的基础上的散布重复序列聚合酶链反应（IRS-PCR）的选择克隆，以确定是否描绘的大片段的SINE探针对应于一个猪染色体或几个染色体的不同片段。该策略得到了双PRINS方法的支持，该方法使用特异于两组不同猪染色体的a-卫星序列的引物。用位于不同猪染色体上的32个标记通过PCR估计每个克隆的基因组保留频率。从152个杂种中，根据细胞遗传学含量和染色体保留频率选择了126个，构建了猪8号染色体辐射杂种图谱。我们对该染色体的初步结果表明，辐射杂交图谱的分辨率比连锁分析获得的分辨率高18倍。
We have developed a panel of 152 whole-genome radiation hybrids by fusing irradiated diploid pig lymphocytes or fibroblasts with recipient hamster permanent cells. The number and size of the porcine chromosome fragments retained in each hybrid clone were checked by fluorescence in situ hybridization with a SINE probe or by primed in situ labeling (PRINS) with SINE-specific primers. A strategy based on the interspersed repetitive sequence polymerase chain reaction (IRS-PCR) was developed for selected clones to determine if the large fragments painted by the SINE probe corresponded to one pig chromosome or to different fragments of several chromosomes. This strategy was buttressed by a double PRINS approach using primers specific for a-satellite sequences of two different groups of swine chromosomes. Genome retention frequency was estimated for each clone by PCR with 32 markers localized on different porcine chromosomes. Of the 152 hybrids produced, 126 were selected on the basis of cytogenetic content and chromosome retention frequency to construct a radiation hybrid map of swine chromosome 8. Our initial results for this chromosome indicate that the resolution of the radiation hybrid map is 18 times higher than that obtained by linkage analysis.