Construction Of Fibroblat Library Of Three Native Livestock And Poultry Breeds In China & Cloning And Sequence Analysis In Gene GPAT Of Beijing Fatty Chicken

Beijing Fatty Chicken, Langshan Chicken and Small-tailed Han Sheep were selected as experimental material. Fibroblast library was constructed through tissue confluency culture and cell freeze technology. Growth dynamics, genetic stability and contamination of of cross breeds etc.were detected through growth curve, chromosome analysis, isoenzyme analysis etc. Cloning and sequence of gene GPAT of Beijing Fatty Chicken was analyzed by cloning technology and software related. The objective of this paper is to supply basis for conserving the genetic resource of the three breeds at cellular level and supply basis for conservation of quality of Beijing Fatty Chicken breed at genetic level. And the study also provides reference for breed resource conservation of other livestock and poultry. Results are as follows:1 The cultured cells appeared as irregular triangle or shuttle, and the monolayer cells growed as radioactive, blaze or helix in shape.2 The prefreezed cell survive rate of Beijing Fatty Chicken, Langshan Chicken and Small-tailed Han Sheep were 97.5%, 96.0%, 97.0% respectively, and the survive rate after recovery were 95.0%, 94.5%, 94.7% respectively.3 The curve was classic "S" type that showed the population doubling time (PDT) was 24 hours since the second day, and cell growth decreased after culture for 5 days.4 Results of chromosomic analysis showed that the chromosome of Beijing Fatty Chicken, Langshan Chicken and Small-tailed Han Sheep were mainly diploid and the individuals of diploid occupied 87.1%, 86.7%, 93.4% in population observed respectively. The chromosomic structure mutation was 1.5%, 0.8%, 1.4% respectively that different from chromosomic morpHology results reported by other researchers.5 The results of analysis on LDH isoenzyme spectrum showed that there were 2 spectrums in Beijing Fatty Chicken and Langshan Chicken, and 5 spectrums in Small-tailed Han Sheep. The result of analysis on MDH isoenzyme spectrum shows that there were 2 spectrums in all the three breeds, but mobility of Small-tailed Han Sheep was significant higher than the 2 poultry breeds.6 Detection for contamination with microbes shows that cultured cells of the 3 breeds appear negative.7 Plasmid, pEGFP-N3, was transfected by lipid body into fibroblast cells of the three breeds, and the study shows that good transfection can be gained after transfection for 10 hours.8 The results of gene GPAT cloning and sequence analysis of Beijing FattyChicken show that GPAT is 953bp in size containing 41.24% (G+C) content. Gene GPAT has 84.63% homology with chicken's and 5 nt inserts hi left flanking after 365bp, but 5 nt is deleted between 423-429 bp and 1nt deletion at 631bp. GPAT of Beijing Fatty Chicken has 81.41% homology with that of human, 3 and 2 nt deletion happen at 243 bp and 326 bp respectively. GPAT of Beijing Fatty Chicken has 78.09% homology with that of mouse and there is 1 nt deletion at 145 bp and 556 bp respectively. 5 Hae sites, 2 Alw26Isites and 3 TaqI sites exist on GPAT of Beijing Fatty Chicken through analysis by Oligo6.0 software.