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 To evaluate the genetic diversity and phylogenetic relationships of Amomun tsao-ko populations in Yunnan. Methods Seven pairs of microsatellite (SSR) primers were used to analyze 24 A. tsao-ko populations; First, GenALEx was used to calculate genetic diversity parameters, PCoA and AMOVA analysis was carried out; NTsys software was then used to draw population clusters map; And finally, the Structure software was used to calculate the best K value. Results The average of Shannon’s diversity index (H) of the 24 A. tsao-ko populations was 0.49, the average of heterozygosity (He) was 0.32, the genetic differentiation coefficient (Fst) was 0.090, and the gene flow (Nm) was 2.930. Eighty-one percent of the genetic differentiation among the 24 populations of A. tsao-ko existed within the population, and only 19% existed among the populations. The genetic identity (I) of the 23 A. tsao-ko populations of yellow flowers was 0.631 8-0.982 4. The genetic distance (D) was in the range of 0.017 7- 0.459 2, while the consistency degree of the A. tsao-ko population of white flower (MG5) and 23 other yellow flowers was 0.369 7-0.609 0. However, cluster analysis showed that A. tsao-ko population of the white flowers and yellow flowers were clearly separated at the genetic distance of 0.49. Structure clustering showed 209 A. tsao-ko resources can be divided into four groups when K value was 4. Conclusion The genetic diversity of A. tsao-ko populations of yellow flowers of Yunnan is higher on average, and the genetic variation is mainly found in population rather than among populations. According to the genotypes, A. tsao-ko of yellow flower and white flower are clearly divided into two categories, and the genetic distance is very far; and the yellow flower of A. tsao-ko is roughly divided into four groups.