Pueraria Efficacy Ingredients Geographic Variation And Genotyping Study

There are abundant resources of pueraria in China,in which the highest content of puerarin is pueraria lobata.However,under different geographical conditions,the efficacy content of puerarin varianced comparatively large,but whether ingredients efficacy Puerarin variation affected by the gene is unclear,to explain pueraria gene variation,study the genotype of it,which is of great significance for its good breeding of pueraria resources.In order to clarify the issue of genetic variation in Pueraria lobata,we carried out the following work,①collected of a total of 24 samples of pueraria from eight different provinces and cities of origins,determined by four DNA barcode gene sequences,study the genotyping of different geographical origin of the Pueraria samples.②Determined the total flavonoids content of pueraria of their roots and content of puerarin in the stem and leaf,studied the geographical variation of ingredients Puerarin efficacy.③Make clear genotyping of puerarin,with a view to providing an excellent seed source Puerarin filter to provide a basis for the molecular evolution Puerarin relationship described preliminarily.The main conclusions are as follows:1.ITS sequence variation Puerarin larger samples of different origin genotype Puerarin more ingredients can not be used to evaluate the efficacyOf the ITS sequences Puerarin,ITS1 and ITS2 sequences,ITS1 sequence,ITS2 sequence,respectively,the cluster analysis results show that the experiment could be the origin of eight different samples Puerarin is divided into 13 genetypes,24 of 12 samples sequence of genotype for the general,non-site variability,and another 12 samples of their ITS sequences occurred in more than one site to the variation between different places and the sequence of the same origin have the phenomenon of cross-clustering,information on the basis of these variations analysis,Puerarin genotype samples from different areas are more ingredients can not be used to evaluate the efficacy. 2.Chloroplast DNA tmL intron sequences without variation,can not be used for genotypingThe use of the tmL intron sequence without mutation sites from different regions,which can not be used for genotyping.3.psbK-psbI sequence variation and geographical distribution can be divided into two different places Puerarin gene types.Using psbK-psbI sequence analysis of this study eight samples Puerarin origin and found that variation associated with the origin,in different places with two Puerarin site variability information,which in the first eight samples and the base 249 base 301 Department of base G,16 samples of these two bases in the Department for T,were named as "TT-type","GG-type."4.trnH-psbA sequence variation and geographical distribution can be divided into two different genotypes Puerarin origin.The use of trnH-psbA sequence analysis of this study eight samples Puerarin origin and found that with the origin of the variation in the first 45 base pairs Department for nine samples of A,15 samples for T,in the 181-191 base nine samples Service There are "ACGTTTTTGA" the existence of paragraph 15 of the sequence of missing samples,respectively,named as "A + non-deletion" and "T + deletion."5.There were significant geographical differences in the content of efficacy ingredients in pueraria.The content of puerarin Flavonoids was in between 9.20～9.65%of different various origin,RSD 0.71%,the difference was not statistically significant,HPLC method used to detect the pueraria lobata root content from the different origin,the content changes from 0.35 to 8.24%range,RSD value of 55%,significantly different; deplete the growth of life factors,determination of the different origin of puerarin in Pueraria lobata leaf content,the content changes from 0.83～4.47%,RSD value of 60%,results show that the difference was extremely significant.6.Puerarin chloroplast gene psbK-psbI and trnH-psbA genotypes has obvious relevance with content,obvious different between them.Using chemical analysis method to study the efficacy of puerarin content components,estimated the puerarin genotyping results,The results showed that different genotypes of Puerarin obvious differences in content,"GG-type" represents the origin of Puerarin content of 2.51%,"TT-type" represents the origin of Puerarin content of 4.55%,"A + non-deletion" represented by the origin of Puerarin content of 2.83%,"T + missing type" represented by the origin of Puerarin content of 4.49%. accordingly screen out high-quality Puerarin genotype,that is,genes psbK-psbI the "TT-type",trnH-psbA gene in the "T + deletion.7.Puerarin chloroplast gene psbK-psbI and trnH-psbA gene is closely related to type and origin.Liaoning Qianshan,Beijing Huairou,Hebei Yixian,Hebei pingli,Henan Lushi for genotype "TT",Shanxi Pinglu,Shaanxi-Pingli,Shaanxi Xunyang for genotype "GG";to Shanxi Pinglu and Henan Lushi as a dividing line,north for the "TT-type", the south as "GG-type",Shanxi Pinglu and Henan Lushi are two heterozygous genotypes area.Shaanxi Pingli which a sample for the "TT-type",different from the other two samples of origin,to be studied further.Liaoning Qianshan,Beijing Huairou,Hebei Yixian,Hebei Pingli,Henan Lushi for "T+deletion",Shanxi Pinglu,Shaanxi-Pingli,Shaanxi Xunyang for "A + non-deletion." In Shanxi Pinglu and Henan Lushi as a dividing line,north to "T + deletion" south to "A + non-deletion",Shanxi Pinglu and Henan Lushi are two heterozygous genotypes area.To sum up,the use of different genotyping germplasm resources Puerarin origin of great significance.Conclusions of this study was to clarify the mechanism of high-quality ingredients to provide the molecular basis of quality;for the screening of fine varieties Puerarin guide Puerarin cultivation,lay the foundation to improve the quality of Chinese medicine.