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Fig. 10 | Reproductive Biology and Endocrinology

Fig. 10

From: Divergent activity of the gonadotropin-releasing hormone receptor gene promoter among genetic lines of pigs is partially conferred by nuclear factor (NF)-kB, specificity protein (SP)1-like and GATA-4 binding sites

Fig. 10

A bp substitution (G → T) located at −845 of the Meishan promoter allows GATA-4 to bind. a Oligonucleotide probes (5’ to 3’) were synthesized containing sequence flanking a naturally occurring point mutation at −845 of the Control/Index (C/I −855/−835) and −843 of the Meishan (M −853/−833) promoter. Underlines within each DNA probe represent the nucleotide substitution between lines and the putative GATA element (TGATAT; Red) identified by sequence analysis is highlighted. b EMSAs were performed by incubating radiolabeled oligonucleotides with nuclear extracts (5 μg) from αT3-1 cells and specificity of DNA-protein interactions was assessed by competition with 50-fold molar excess of homologous or heterologous unlabeled DNA (specific complexes indicated by arrows). Additionally, the DNA-protein complex was challenged by competition with 50-fold molar excess of unlabeled oligonucleotides containing consensus binding sequences for GR, NF-1 and GATA. c To determine the specific factor(s) comprising the Meishan-specific complex, nuclear extracts were also incubated with antibodies directed against GATA-1, -2 and -4 or an equal mass of rabbit IgG (supershift indicated by arrows)

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Reproductive Biology and Endocrinology

ISSN: 1477-7827

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