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Construction and Characterization of Two Bacterial Artificial Chromosome Libraries of Zhikong Scallop, Chlamys farreri Jones et Preston, and Identification of BAC Clones Containing the Genes Involved in Its Innate Immune System

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Abstract

Large-insert bacterial artificial chromosome (BAC) libraries are necessary for advanced genetics and genomics research. To facilitate gene cloning and characterization, genome analysis, and physical mapping of scallop, two BAC libraries were constructed from nuclear DNA of Zhikong scallop, Chlamys farreri Jones et Preston. The libraries were constructed in the BamHI and MboI sites of the vector pECBAC1, respectively. The BamHI library consists of 73,728 clones, and approximately 99% of the clones contain scallop nuclear DNA inserts with an average size of 110 kb, covering 8.0× haploid genome equivalents. Similarly, the MboI library consists of 7680 clones, with an average insert of 145 kb and no insert-empty clones, thus providing a genome coverage of 1.1×. The combined libraries collectively contain a total of 81,408 BAC clones arrayed in 212 384-well microtiter plates, representing 9.1× haploid genome equivalents and having a probability of greater than 99% of discovering at least one positive clone with a single-copy sequence. High-density clone filters prepared from a subset of the two libraries were screened with nine pairs of Overgos designed from the cDNA or DNA sequences of six genes involved in the innate immune system of mollusks. Positive clones were identified for every gene, with an average of 5.3 BAC clones per gene probe. These results suggest that the two scallop BAC libraries provide useful tools for gene cloning, genome physical mapping, and large-scale sequencing in the species.

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Acknowledgements

This project is supported by Key Program of National Natural Science Foundation of China (No. 30730071), Major State Basic Research Development Program of China (2006CB101804), and National High-Tech Research and Development Program of China (2007AA09Z430). The authors thank Prof. Linsheng Song, Prof. Bin Liu, Dr. Bing Wang, Dr. Jiquan Zhang, Miss Fan He, and other laboratory members for technical advice and assistance.

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Correspondence to Jianhai Xiang.

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Yang Zhang and Xiaojun Zhang contributed equally to the article.

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Zhang, Y., Zhang, X., Scheuring, C.F. et al. Construction and Characterization of Two Bacterial Artificial Chromosome Libraries of Zhikong Scallop, Chlamys farreri Jones et Preston, and Identification of BAC Clones Containing the Genes Involved in Its Innate Immune System. Mar Biotechnol 10, 358–365 (2008). https://doi.org/10.1007/s10126-007-9071-8

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  • DOI: https://doi.org/10.1007/s10126-007-9071-8

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