Document Detail

Investigating Transgene Integration and Organization in Cotton (Gossypium hirsutum L.) Genome.
MedLine Citation:
PMID:  23143486     Owner:  NLM     Status:  In-Data-Review    
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.
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    
Cotton Research Center, Shandong Academy of Agricultural Sciences, Jinan, Shandong, P.R. China.
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