Wang W M, Jiang G H, Wang S Q, Zhu L H, Zhai W X
Institute of Genetics, Plant Biotechnology Laboratory, Chinese Academy of Sciences, Beijing 100101, China.
Yi Chuan Xue Bao. 2001;28(2):120-8.
A BAC library for IRBB56, an accession pyramiding Xa4, xa5 and xa13 three bacterial blight resistance genes, was constructed. The library contains 55,296 clones with an average insert size of 132 kb. Based on a haploid genome size of 450 Mb, the coverage of the library was about 14 genome equivalents that make it one of the most comprehensive BAC libraries available in rice and provide 99.99% possibility to isolate any interested rice genes or sequences in the library. To determine the representation of organelle DNA homologues in the library, the library was screened with three different chloroplast genes and four mitochondrial genes, respectively. Results from this screening showed that less than 1% of clones in the library contain organelle genomic DNA homologues. Then, DNA markers on three different chromosomes linked to Xa4, xa5, and xa13, and a PCR fragment of rice UROD gene, were used to screen the library resulting in a range of 11-106 hits that will promote the isolation of these genes. The deep coverage and the large insert size of the library will facilitate physical mapping, isolation, and cloning of rice genes.
构建了一个用于IRBB56的细菌人工染色体（BAC）文库，IRBB56聚合了抗白叶枯病基因Xa4、xa5和xa13。该文库包含55,296个克隆，平均插入片段大小为132 kb。基于单倍体基因组大小450 Mb，该文库的覆盖度约为14个基因组当量，使其成为水稻中最全面的BAC文库之一，并提供了99.99%的可能性来分离文库中任何感兴趣的水稻基因或序列。为了确定文库中细胞器DNA同源物的代表性，分别用三个不同的叶绿体基因和四个线粒体基因对文库进行筛选。筛选结果表明，文库中不到1%的克隆含有细胞器基因组DNA同源物。然后，使用与Xa4、xa5和xa13连锁的三个不同染色体上的DNA标记以及水稻尿卟啉原脱羧酶（UROD）基因的一个PCR片段对文库进行筛选，得到了11 - 106个阳性克隆，这将有助于这些基因的分离。该文库的高覆盖度和大插入片段大小将有助于水稻基因的物理图谱构建、分离和克隆。
