GeXP Multiplex PCR Technology To Identify The Species Of Deer

Deer and its products were considered to be superb food and health products since ancient times.Two of authoritative pharmacopeia < Shennong Bencao> and <Compendium of Mate ria Medica> both have recorded its supetior function,such as warming the kidney and strength-ening yang,replenishing essence and blood and other effects.In 2000,the sika deer and the red deer were included in the "People's Republic of China Pharmacopoeia"(a)as genuine her bs,Thereafter the development and application of high-value deer food and drug health care products gradually increased,followed by various types of deer products and its processed products at home and abroad getting varying degrees of adulteration doping,and its wide range of doping,adulteration is more diversified.There are three typical types of adulteration: shoddy,adulterated,completely pseudo-type,each caused interference and threatst o the consumer's economy,security,and special religious beliefs.In addition,in order to prevent white deer deer and other national key protected wild deer from being illegally killed,the need for deer animal protection and regulatory means put forward higher requirements,thus to establish a scientific,specific,accurate,high-throughput deer inter-species identification Method is very necessary.In this study,research based on the technology of GeXP multiplex PCR was used to identify 6 species of deer,including Cervus elphus,Rangifer tarandus,Cervus nippon,Elaphurus davidianus,dama dama,Cervus albirostris.The main research work and results as follows.1.Based on the results of DNAMAN on the nucleotide sequence of D-loop,cytb,16 S rRNA gene of deer species,we designed and screened 150 pairs of specific primers of 6 species of deer,and finally confirmed The length of the amplified product fragment: Cervus elphus was 283 bp,the reindeer was 107 bp,the Elaphurus davidianus was 157 bp,the dama dama was 167 bp,and the white deer deer was 187 bp,Cervus nippon / elphus was 316 bp.All of the specificity of primers were analyzed by agarose gel electrophoresis and GeXP analysis.The results shows effective only for the 6 species of deers.2.The single factor analysis method was used to optimize the cross-specificity of the three types of DNA polymerase,6 groups of different primer annealing temperature and three groups of different primers concentration ratios in GeXP multiplex PCR system.By repeated experiments we determined the ideal parameters adapting to the study,and a specific,sensitive,clear detection system was established.3.Two kinds of GeXP multiplex PCR system were established.One was to the red deer products for the Cervus elphus,Rangifer tarandus,Cervus nippon,Elaphurus davidianus,dama dama,Cervus albirostris.The other was to the red deer or cervus nippon products.The system was able to accurately identify the composition of all the 6 species of deer but sika deer,and the sika deer can be distinguished under the condition of no red deer.While the mixture with red deer fails.4.16 commercial samples were collected and tested by the GeXP multiplex PCR system.The results showed that the method was sensitive and specific,which could be effective and reliable for deer species differentiation,specific deer species and deer products in deer products.Presently,the reports of the use of GeXP multiplex PCR technology at home and abroad for gene expression analysis and pathogen detection research has already been very much,however deer-species distinction never.What's more,researches about making a multiplex identification to species of deer in only one experiment is also in a great shortage no matter at home or abroad.The above all as the research innovation and significant point.