Investigating Transgene Integration and Organization in Cotton (Gossypium hirsutum L.) Genome

文献类型： 外文期刊

第一作者： Zhang, Jun

作者： Zhang, Jun;Hong, Yan

作者机构：

关键词： Transgenic cotton; Transgene/plant genome junctions; Transgenic structure; Transgene repeats; Vector backbone; Conventional PCR; Genomic Southern blot hybridization; Inverse PCR

期刊名称：TRANSGENIC COTTON, 2 EDITION: Methods and Protocols

ISSN： 1064-3745

年卷期： 2019 年 1902 卷

页码：

收录情况： SCI

摘要： In this chapter, we present detailed experimental procedures for investigating integration patterns of transgenes in cotton genome. We use conventional PCR and genomic Southern blot hybridization to characterize integration of T-DNA components and vector backbone fragments. For multiple-copy insertions into the same site (complex loci), transgene/transgene junctions (including canonical and truncated T-DNA and transgene involved vector backbone sequences) are characterized by PCR and sequencing. Inverse PCR and sequencing are used to characterize transgene/cotton genome junctions. Distribution of T-DNA insertion in cotton genome is evaluated by analysis of transgene flanking sequences. The pre-insertion sites can also be cloned and sequenced (based on the flanking sequences) for survey of genomic structure changes brought by transgene integration by comparing a pre-insertion site with corresponding transgene/plant junctions.

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