The Polymorphisms Of Prion Gene Family In Wenzhou Buffalo

The transmissible spongiform of encephalopathy on cattle (mad cowdisease, BSE) is a neurodegenerative disease that happened in human and mammals.Human can be infected by the cattle through oral, and the mortality rate was up to100%. In addition to control the various feed, food, blood products, medicalequipment and other ways from cattle, selection for animals with resistant gene for acertain disease is a cutting-edge technology as an important means of resistantanimal breeding. The previous studies had proved that the genetypes of PRNP (oneof the prion gene family) had important effect on the resistance to scrapie in sheep,and another two members of the prion protein gene families PRND and SPRNincreasing focused people’s attention and research. As important candidate genes,the insertion and deletion polymorphisms of the23bp sequence in the promoterregion and the12bp sequence in intron1of PRNP have a great effect on BSEsusceptibility or resistance, both of them can regulate the expression level of PrP bythe combination or separation with the transcriptional regulation factors, and theninfluence the resistance to BSE.In this study, Wenzhou buffalo was choosed for the research material whichnever been found cases of BSE as well as other buffalo breeds, and68Holstein cowsamples and reported BSE cattles as control. PCR and the denatured polyacrylamidegel electrophoresis were used to determine the insertion/deletion polymorphisms andtheir frequencies within the95WenZhou buffalo samples. Finally, the resultsshowed that the allele frequencies and genotype frequencies had significantlydifference between WenZhou buffalo and other breeds of cattle including Holestincow and a beef cattle breed, which predicted the buffalo had disease resistance toBSE. Detection and analysis results showed that the WenZhou buffaloes all hadinsertion/insertion homozygous genotypes at23/12bp polymorphic loci, which werein consistent with the conclusion that reported in other water buffaloes. Thefrequency of insertion allele in water buffalo reported was more than90%. Thesignificant difference (p<0.01) results indicated that the WenZhou buffalo have astronger resistance to BSE than Holstein cow.Secondly, there is a highly conservative octapeptide repeat area (that is the polymorphism of24bp sequence) in the coding region of PRNP, whose repeatnumber and the resistance of BSE had an negative correlation, and the repeat numberwas6in most of the individuals and the other was5(the average frequencies are0.894and0.106respectively). In this study, the number of octapeptide repeat wasdetected to estimate whether has a significant difference between WenZhou buffaloand the other cattle previously reported. The results showed that WenZhou buffaloand Holstein cow have no significant differences (p>0.05) in the polymorphism of24bp sequence. It proved that there may be have no significant relationship betweenthe strong disease resistance and the number of octapeptide repeat. PCR andsingle-strand conformation polymorphism and PAGE methods were used to detectthe SNPs located in β2-α2-loop of PrP which probably have an important effect onresistance, and one SNP site at600th nucleotide was acquired which didn’t result inthe change of the amino acid residue.Furthermore, the polymorphisms of PRND encoding Dopple protein, anothermember of prion gene family, was studied in order to compare the difference ofbuffalo and other cattle. By SSCP-PAGE analysis, some differences of the PRNDgene were detected between buffalo and cow, and different nucleotides resulted inamino acid residues change, but there only had4SNPs between WenZhou buffaloand Bubalus bubalis, a river buffalo in India, two of which cause the amino acidschange, and they were both heterozygous mutations. Especially the specificnucleotide sequence that exist in buffalo rather than Bubalus bubalis. The SNPsfound between WenZhou buffalo may offer a new possibility for further diseaseresistance study.