ABSTRACT
ObjectiveTo identify the molecular biology of various species of Tibetan Codonopsis plants based on internal transcribed spacer（ITS）2 and psbA-trnH sequence barcode technology. MethodThe genomic DNA of 28 Tibetan Codonopsis plant samples from four species （Codonopsis canescens，C. foetens subsp. nervosa，C. pilosula， and C. thalictrifolia var. mollis） were extracted，and the ITS2 and psbA-trnH sequences were amplified and sequenced. The related sequences of 81 Tibetan Codonopsis plant samples belonging to 15 species were downloaded from GenBank， and MEGA 6.0 was used for sequence comparison and mutation site analysis. The GC content and genetic distance within and between species were calculated. Additionally， phylogenetic trees were constructed by maximum likelihood （ML） method， neighbor-joining （NJ） method，and unweighted pair-group method with arithmetic means （UPGMA） . ResultAccording to the mutation site，C. canescens， C. pilosula，C. pilosula subsp. tangshen， C. pilosula var. modesta，C. bhutanica，C. clematidea，C. lanceolata，C. subglobosa and C. foetens were distinguished. In the phylogenetic trees，the optimal clustering effects for ITS2 and psbA-trnH sequences were obtained using the ML method and the UPGMA method， respectively， and 12 species were effectively clustered. ConclusionITS2 and psbA-trnH sequences have a high identification rate for species of single origin，but there are still some limitations in identifying variants and original variants. This study provides basis for the identification of affinity relationship and clinical safety of Tibetan Codonopsis plants.
ABSTRACT
Objective: To screen and evaluate DNA barcoding of Amomun tsao-ko populations in Yunnan. Methods: ITS, psbA-trnH, matK, rbcL, and ycf1 sequences were screened and evaluated using A. tsao-ko as samples. The samples of A. tsao-ko population were amplified and sequenced. The sequences were spliced with Genestar, and then processed with Mega for data processing. And A. tsao-ko diversity and identification were analyzed and discussed. Results: The length of the amplified fragments of primers ITS5 and ITS4 was approximately 520 bp; The length of the amplified fragments of the primers rbcLa-F and rbcLa-R was approximately 498 bp; The length of the amplified fragments of the primers ycf1-bF and ycf1-bR was approximately 800 bp; The length of the amplified fragments of the primers psbA-trnH-1F and psbA-trnH-1R was approximately 400 bp; The length of the amplified fragments of the primers matK-2F and matK-2R was approximately 470 bp. The success rate of amplification and sequencing was high, and most of the results were available. By analyzing the amplification results of ITS, psbA-trnH, matK and ycf1 sequences of A. tsao-ko, A. tsao-ko and other Amomum genus plants can be clearly distinguished; All samples of the ITS sequence were divided into MG5 white flower A. tsao-ko population and other populations; All samples of the psbA-trnH sequence were divided into MG5 white flower A. tsao-ko population, MG6 yellow flower A. tsao-ko population and other populations; All samples of the matK sequence were divided into MG6 A. tsao-ko population and other populations. The MG5 white flower A. tsao-ko sample failed to be amplified; All samples of the ycf1 sequence were divided into the MG6 yellow flower A. tsao-ko population and other populations, and the MG5 white flower A. tsao-ko population was clustered with the other 22 A. tsao-ko populations; The amplification of rbcL sequence was consistent for all samples. Conclusion: The ITS, matK, psbA-trnH and ycf1 sequences can accurately distinguish A. tsao-ko from other plants of Amomum genus; The sequence site variations were found in matK, psbA-trnH and ycf1 sequences of MG6. This research has contributed to the selection and breeding of A. tsao-ko varieties. ITS and psbA-trnHsequences can distinguish yellow flower and white flower of A. tsao-ko; There is no variation in the rbcL sequence of all samples of white and yellow flowers of A. tsao-ko, and Amomum tsao-ko and other plants of Amomum genus cannot be identified with the rbcL sequence, which can be discarded.
ABSTRACT
Objective System evolution relationship and molecular identification method of the germplasm resources of Lycoris aurea from different regions was analyzed based on the sequence of psbA-trnH chloroplast gene. Methods DNA samples of 52 L. aurea populations were extracted from 15 provinces or cities in China. The psbA-trnH sequences of the populations were amplified by PCR, and the purified PCR products were sequenced and analyzed by Mega 5.0 software etc. Results The length of psbA-trnH sequences were 544-656 bp, and GC content of them was 35.8%-37.0%, and the genetic distances among the populations were 0-0.009 47. There were 33 variable (polymorphic) sites, including nine parsimony informative sites and 18 singleton variable sites and six insertion/deletion gaps. Ten haplotypes (H) were identified. Values of haplotype diversity (Hd) and nucleotide diversity (π) were 0.749 and 0.002 63, respectively. The genetic diversity of the populations of L. aurea were very high. In the maximum parsimony phylogenetic tree, 52 populations of L. aurea were clustered into four branches, which was almost consistent with their geographical distributions. Conclusion The genetic variation of L. aurea populations from different regions is significant and the psbA-trnH sequence could be used as a molecular evidence for identifying the germplasm resources of L. aurea from different regions. There is very obvious regional characteristics in evolution for germplasm resources of L. aurea in China.