Document Detail


Investigating Transgene Integration and Organization in Cotton (Gossypium hirsutum L.) Genome.
MedLine Citation:
PMID:  23143486     Owner:  NLM     Status:  In-Data-Review    
Abstract/OtherAbstract:
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 (see Note 1) and sequencing is 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.
Authors:
Jun Zhang; Yan Hong
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Methods in molecular biology (Clifton, N.J.)     Volume:  958     ISSN:  1940-6029     ISO Abbreviation:  Methods Mol. Biol.     Publication Date:  2013  
Date Detail:
Created Date:  2012-11-12     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  9214969     Medline TA:  Methods Mol Biol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  95-107     Citation Subset:  IM    
Affiliation:
Cotton Research Center, Shandong Academy of Agricultural Sciences, Jinan, Shandong, P.R. China.
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