Study On The Molecular Identification Of Antler And The Difference Of Antler Protein

Objective:To establish an efficient method for extracting DNA from velvet antler and to establish a new PCR identification method based on the CO? sequence to quickly identify sika deer,red deer and reindeer antler;optimize the optimal process of antler protein extraction and perform antler and red antler Proteomics Research.Methods:Four kinds of antler genomic DNA extraction methods were investigated with reference to the 2015 edition of the Pharmacopoeia of the People's Republic of China.By comparing the sequence differences of the CO? gene,designing identification primers for velvet antler,red antler and reindeer antler based on SNP identification sites,and examining the PCR reaction conditions Response surface methodology was used to optimize the extraction process of velvet antler protein;the Proteomic analysis of velvet antler and red antler protein was performed by searching the Uniprot database,and differentially expressed proteins were identified by bioinformatics software.Results:Results: Using Tiangen kit,70 mg of pilose antler powder,56 ? water bath temperature,and 6 h of water bath time,the genomic DNA extraction of pilose antler was the best.The two-site specific PCR identification system based on the CO? gene sequence developed in this study can finally Sika deer and red deer specific electrophoresis bands were observed at 294 bp,while reindeer specific bands were observed at 514 bp.No other fakes and negative controls had bands;optimized ultrasonic fragmentation method for extracting velvet protein conditions: pH 7.6,Ultrasound time was 24 minutes,liquid-to-liquid ratio was18:1(mL/g),lysis time was 60 minutes,and the extraction rate reached 3.82%.After proteomics research,36 proteins were highly expressed in velvet antler,and 144 proteins were expressed in velvet Medium and high expression,GO and KEGG analysis showed that differentially expressed proteins are involved in the regulation of several pathways,including the PI3K-Akt signaling pathway,tissue heat generation,and oxidative phosphorylation.Conclusion:The method of extracting antler DNA from the Tiangen kit optimized in this study is simple and convenient,and the DNA concentration and quality are good.Theestablished two-site PCR method for identifying sika deer,red deer,and reindeer other fake antler is simple and accurate;The ultrasonic extraction method of pilose antler protein is stable and effective,laying a foundation for subsequent proteomics research.Proteomics studies revealed the differential expression of proteins in velvet antler and red antler in terms of cellular components,biological metabolism,molecular functions and pathways.