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Construction of BIBAC and BAC libraries from a variety of organisms for advanced genomics research

Nature Protocols volume 7, pages 479–499 (2012)Cite this article

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Abstract

Large-insert BAC (bacterial artificial chromosome) and BIBAC (binary BAC) libraries are essential for modern genomics research for all organisms. We helped pioneer the BAC and BIBAC technologies, and by using them we have constructed hundreds of BAC and BIBAC libraries for different species of plants, animals, marine animals, insects, algae and microbes. These libraries have been used globally for different aspects of genomics research. Here we describe the procedure with the latest improvements that we have made and used for construction of BIBAC libraries. The procedure includes the preparation of BIBAC vectors, the preparation of clonable fragments of the desired size from the source DNA, the construction and transformation of BIBACs and, finally, the characterization and assembly of BIBAC libraries. We also specify the modifications necessary for construction of BAC libraries using the protocol. The entire protocol takes ∼7 d.

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Figure 1: An overview of the procedure for construction of BIBAC libraries.
Figure 2: Demonstration of BIBAC as well as BAC library construction.
Figure 3: BIBACs and BACs constructed using this protocol.

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Acknowledgements

We thank the former members of H.-B.Z.'s laboratory who have used the methods described in this article for their research and thus might have directly or indirectly contributed to the procedures. This work is supported in part by Research Grant Award no. IS-4427-11C from BARD, the United States-Israel Binational Agricultural Research and Development Fund (H.-B.Z.) and grants (124329-85360; 124475-85360) from the Texas AgriLife Research Cotton and Monocot Crop Improvement Program (H.-B.Z.).

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Author notes

  1. Cheng-Cang Wu, Jennifer J Dong & Yaning Li

    Present address: Present addresses: Lucigen Corporation, Middleton, Wisconsin, USA (C.-C.W.); Department of Microbial and Molecular Pathogenesis, Texas A&M Health Science Center, College of Medicine, College Station, Texas, USA (J.J.D.); and College of Plant Protection, Agricultural University of Hebei, Biological Control Center of Plant Disease and Plant Pests of Hebei Province, Hebei, China (Y.L.).,

Authors and Affiliations

  1. Department of Soil and Crop Sciences, Texas A&M University, College Station, Texas, USA

    Hong-Bin Zhang, Chantel F Scheuring, Meiping Zhang, Yang Zhang, Cheng-Cang Wu, Jennifer J Dong & Yaning Li

  2. College of Life Science, Jilin Agricultural University, Changchun, Jilin, China

    Meiping Zhang

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Contributions

H.-B.Z., C.F.S., M.Z., Y.Z., C.-C.W., J.J.D. and Y.L. conducted the experiments on different species with the protocols presented here, and improved and extended the original protocol through these experiments. Furthermore, H.-B.Z. developed the original concept of the protocols and the original protocols, designed the experiments and wrote the manuscript.

Corresponding author

Correspondence to Hong-Bin Zhang.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Figure 1

BAC and BIBAC vectors widely used in the construction of BIBAC and BAC libraries. (a) pECBAC1 (ref. 63) and (b) pBeloBAC11 (ref. 64) are both BAC vectors, whereas (c) pCLD04541 (561, d) BIBAC2 (462) and (e) pYLTAC7 (ref. 6) are BIBAC vectors. The cloning sites of restriction enzyme(s), the gene marker for selection of transformants (CMR, chloramphenicol; TetR, tetracycline; KmR, kanamycin) and the gene marker for selection of clones (Lac Z or Sac B) containing inserts or recombinant clones are highlighted or in bold. The figure was from Ref. 35 with permission from its publisher, Wiley-VCH. (TIFF 5426 kb)

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Zhang, HB., Scheuring, C., Zhang, M. et al. Construction of BIBAC and BAC libraries from a variety of organisms for advanced genomics research. Nat Protoc 7, 479–499 (2012). https://doi.org/10.1038/nprot.2011.456

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