Screening, Identification And Characterization Of Differentially Expressed Genes In Muscle Tissue Between F₁ Of Meishan×Largewhite And Largewhite

Differential gene expression is mainly responsible for heterosis between parents and progeny. Based on suppression subtractive hybridization(SSH), this study, aimed at laying groundwork for isolating and identifying genes related with heterosis as well as exploring the molecular mechanism of heterosis at the level of gene expression, generated the forward- and reverse- subtractive cDNA libraries containing some differentially expressed fragments between Longissimus Dor si of Largewhite and FI of Meishan X Largewhite pigs. The findings are as follows:(1) Both of the two subtractive cDNA libraries were subject to the determination of subtraction efficiency that proved to be approximately 215- and 210-fold indicated by housekeeping gene 3 -actin as an index and this suggests that the libraries are well constructed for subsequent study.(2) A total of 1,500 clones in the plasmid libraries was obtained after the cDNA fragments were ligated to pGEM-T vector and then transformed to E.coli JM109 by picking up the white colonies at random.(3) PCR analysis of the total of 700 and 623 positive clones derived from the corresponding plasmid libraries demonstrates that the cDNA inserts range in size largely from 150~600bp.(4) 39,21 differentially expressed fragments were determined out of 200, 150 clones in FI and Largewhite subtractive libraries, respectively.(5) Sequencing and analyzing by BLASTN, BLASTX the 39 differentially expressed fragments in FI subtractive library indicate that they are from 6 genes , i.e., Homo sapiens spliceosomal protein SAP155 -. Sus scrofa mRNA for glyceraldehyde-3-phosphate dehydrogenase (G3PDH)-. Homo sapiens myosin binding protein H^ Sus scrofa mRNA for myosin light chain N Sus scrofa integrin beta-1 subunit (CD29) mRNA^ Sus scrofa cytochrome oxidase subunit I.(6) Sequencing and analyzing by BLASTNU BLASTX the 21 differentially expressed fragments in Largewhite subtractive library indicate that they are from 1 genes, i.e., mRNA for glyceraldehyde-3-phosphate dehydrogenase(G3PDH) in Sus scrofa.(7) Integrin beta-1 subunit (CD29) is verified as differentially expressed gene in FI by RT-PCR.