Research On The Differentially-expressed Genes Of Muscle Tissue In Double-muscled Large White Pig

The percentage of lean is one of the most important traits in livestock. In recent years, more and more researchers are interested in the Large White Pigs with double-muscled hips because of their high lean rates. In this research, we used some molecular methods to investigate the relevant genes, which had great effects on the double-muscled hips or high lean rates, in order to accelerate the speed of double-muscled pig breeding.Stastic analysis of four pares of full-twins showed that there was significant difference (|Pr > F| =0.0319) between double-muscled pig (DM) and non-double-muscled (NDM) one, and the bodyweight was also significant difference at 0.10 level, but there was no difference in body length, body height and breast.We firstly constructed two RNA pools were from the full sibs of DM and NDM individually and built up subtracted cDNA library by suppression subtracted hybridization (SSH) further. The subtracted library had 768 clones including 686 effective clones, which percentage was about 90% of total clones. We have gotten 587 high quality sequences by sequencing, 69 of which were not found identical sequences with EST data of Genbank by BLAST procedure. Considered no identity found with EST data or the sequences were in 3' non-translation region with great changes, we selected 11 clones as new sequences in our research.With the radioactive isotope method, subtractive cDNA library was screened and the results were analyzed by image analyzing system. There were 16 clones with the signal strength of forward probes twice higher than that of reverse ones in both results of subtractive cDNA probes and the reverse transcription probes. And there were 33 clones with the signal strength of forward probes twice higher than that of reverse ones only in results of subtractive cDNA library.The results of sequencing and differential screening showed that there were two differentiated expression ESTs in our research: one was new, and the other had been reported but its function was still unknown; and there were many other differentiated expression genes associated closely with the myogenesis such as cardiac a-tropomyosin, MHC 2b, MHC 2x, LDH-A, RYR1, and IGFBP-7. In themeantime, the results of Northern Blotting showed that the mRNA expression level of MHC 2x and MHC 2b in DM pigs were obviously higher than that of NDM pigs. The differentiated expression features of MHC 2x and MHC 2b suggested that DM pigs have a higher proportion of white muscle fibers than red muscle fibers, which also indirectly proved by the differential expression of LDH-A, because MHC family is an important determinant parameters of muscle fibers such as MHC 2b and MHC 2x, which all determines the white muscle fiber type.The results of quantitative PCR showed that the expression levels of RYR1, CAMK2G and IGFBP7 in DM RNA pool were higher than that of NDM RNA pool in DM pigs, respectively. So did No. 695, No. 882 and No. 480 clone. The up-regulated expression of these genes might be closely related to the DM trait.In summary, we concluded that the reasons of DM pigs had more developed muscles in buttocks than NDM pigs might be the former had a higher proportion of white muscle fibers, which are in larger diameters. Moreover, cardiac a-tropomyosin RYR1 IGFBP-7 CAMK2G genes and the genes of No.202, No.321 No.695 as well as No.292 were all the potential candidate genes associated to the DM trait in pigs.