Establishment Of Transgenic Bovine Gene Chip Detection Method

Gene technology is a "double-edged sword" for humanity in the economic and social benefits, but also caused many problems.The GMOs were advantage with survivability and competitive power as a result of being inserted into valuable genes,it may impact on the biodiversity,eco-system and human development,so the detection of GMOs is necessary in spite of the emergence of GMOs'potential risk occur many years latter. With the development of transgenic technology, the commercialization of transgenic animals will gradually expand the scope for possible trouble, the need for scientific testing. The tools applied for GMO testing are primarily ptotein based(mainly immunological)assays and DNA based assays (mainly applying the polymerase chain reaction [PCR]technology),which mostly based on the sequential detection of one target at a time, or on a limited multiplexing, allowing only a few targets to be analysed at once, no longer meet the testing requirements.As a result,the high-througput assays such as microarray have also been proposed,owe its superiority such as high-throughput,micromation and specificity fitting for GMOs detection.A noval multiplex-PCR coupled with oligonucleotide microarray was established for detection of genetically modified bevine. Eight pairs of specific primers and Sixteen specific probes were designed based on the sequences of bovine mtDNA 16s rRNA gene (BOS mtDNA16s rRNA, BOS), Ovis aries mtDNA12s rRNA gene(Ovis mtDNA12s rRNA), Sus cristatus mtDNA NADH5(Sus NADH5), neomycin phosphotrAnsfase (NPTâ…¡), enhanced green fluorescent protein(EGFP), Human Lactoferrin gene(hLF), Human a-Lactalbumin gene(ha-LALBA) and human Lysozyme gene(hLYZ). The method was optimized by the ratio of sense and antisense primers, hybrid system, washing conditions. The sensitivity and specificity of the method were evaluated with Recombinant Human Lactoferrin Transgenic Cloned cow samples, Recombinant Huma a-LALBA Cloned cow samples,Recombinant Human Lysozyme Cloned cow samples and other genetically modified animal samples, the results showed that the method was very specific and sensitive, It's dectction limit is 1pg-A noval multiplex-PCR coupled with oligonucleotide microarray was established not only to detect detect bovine,Ovis aries,and sus derived materials,As well as screen genetically modified bovine, but also to identify the exogenous genes.this study could make the technical reserve ready for the detection of the genetically modified animals.