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Isolation of Putative in vivo Hoxc8 Downstream Target Genes Using ChIP-Cloning Method

 Hyun Joo Chung  ;  Myengmo Kang  ;  Myoung Hee Kim 
 Journal of Experimental & Biomedcal Science (대한의생명과학회지), Vol.14(1) : 47-53, 2008 
Journal Title
 Journal of Experimental & Biomedcal Science (대한의생명과학회지) 
Issue Date
Hoxc8 ; ChIp-Cloning ; Hoxc8 downstream target gene ; Mouse embryo
Hox genes are known to be transcription factors controlling vertebrate pattern formation along the anteroposterior body axis by regulating many target gene expressions during vertebrate embryogenesis. In order to isolate in vivo Hox responsive target genes, ChIP-cloning technique has been applied using Hoxc8 antibody. Here murine embryo of day 11.5 post coitum (E11.5) highly expressing Hoxc8 gene was used after removing head and tail portions where Hoxc8 is rarely expressing. After fixation with formaldehyde, the chromatin DNAs harboring bound proteins were isolated. After sonication, about 0.5- to 1 Kb chromatin DNAs were immunoprecipitated with anti Hoxc8 antibody. After removing the bound proteins with proteinase K, DNAs were isolated, cloned into the pBluescsript II SK vector, and then sequenced. Total 33 random clones sequenced were anlalyzed to be located at 12 different genomic regions. Among these, 8 turned out to be introns and 4 were intergenic regions localized in random chromosomes. The base composition of total cloned genomic sequences (6608 bp) were AT-rich, i.e., 40% GC. When the Hoxc8 core binding sites, such as TAAT, ATTA, TTAT, and ATAA were analyzed total number of 55, 45, 54, and 55 were found, respectively, which are than twice as many as expected number of 26. Although this in silico analysis does not mean that the ChIP-cloned sequence is real Hoxc8 regulatory element in vivo, these results strongly imply that the DNA fragments cloned through chromatin immunoprecipitation could be very much likely the putative Hoxc8 downstream target genes.
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1. College of Medicine (의과대학) > Dept. of Anatomy (해부학교실) > 1. Journal Papers
Yonsei Authors
Kang, Myengmo(강명모)
Kim, Myoung Hee(김명희) ORCID logo https://orcid.org/0000-0001-5652-1452
Chung, Huyun Joo(정현주)
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