Studies On Conservation Genetics Of Camellia Pingguoensis

Camellia pingguoensis D.Fang belongs to Camellia?Theaceae?,including Camellia pingguoensis D.Fang var.pingguoensis and the variety Camellia pingguoensis D.Fang var.Terminalis?J.Y.Liang et Z.M.Su?T.L.Ming et W.J.Zhang.C.pingguoensis is an evergreen shrub that grows in the karst area.C.pingguoensis D.Fang var.pingguoensis is distributed in Pingguo County and Tiandong County of Guangxi,C.pingguoensis D.Fang var.terminalis is only distributed in Tiandeng County of Guangxi.The distribution of C.pingguoensis is very narrow,the wild populations have been illegally transplanted and are now classified as endangered species.We collected a total of 328 individuals from eleven natural populations,seven of which were C.pingguoensis D.Fang var.Pingguoensis from Pingguo County and Tiandong County,and the remaining four populations were Camellia pingguoensis D.Fang var.terminalis from the Tiandeng County.The collected populations covers the entire distributional area of C.pingguoensis.The collected samples were analyzed based on eight pairs of microsatellite primers,a single copy nuclear gene PAL?phenylalanine ammonia-lyase?sequence and a chloroplast small single copy region?SSC?to study the genetic diversity and structure of C.pingguoensis,and protection strategies are propose based on the results.The main findings are as follows:1)Based on microsatellite markers,the results show that C.pingguoensis has a moderate level of genetic diversity.At the overall level,the average number of alleles?N_A?of eleven populations ranged from 2.6 to 6.6 with an average value of 4.6;the number of effective alleles?N_E?ranged from 1.871 to 3.386 with an average value of 2.714;The observed heterozygosity?Ho?is between 0.279 and 0.645 with an average value of 0.48;the expected heterozygosity?He?is between 0.26 and 0.623 with an average value of 0.502.At the subspecies level,the average allele?N_A?of C.pingguoensis D.Fang var.Pingguoensis ranged from 4.5 to 6.6 with an average value of 5.4;the number of effective alleles?N_E?ranged from 2.11 to 3.505 with an average value of2.995;The observed heterozygosity?Ho?ranged from 0.413 to 0.645 with an average value of0.512;The expected heterozygosity?He?ranged from 0.382 to 0.623 with an average value of0.557;The average allele?N_A?of C.pingguoensis D.Fang var.Terminalis is between 2.6 and 4.3with an average value 3.3;the effective allele?N_E?ranged from 1.871 to 2.621 with an average value of 2.221;the observed heterozygosity?Ho?ranged from 0.279 to 0.565 with an average value of 0.424.The expected heterozygosity?He?is between 0.26 and 0.557 with an average value of 0.407.Analysis based on single copy nuclear gene?PAL?and chloroplast fragment?SSC?also showed that C.pingguoensis had moderate levels of genetic diversity?cpDNA,h=0.884,?=0.00082;PAL,h=0.839,?=0.0043?.The sequence of PAL is 662 bp in length and contains nineteen simple polymorphic loci and six single nucleotide variant loci.Among them,haplotypes H1 and H2 are the shared haplotypes of C.pingguoensis D.Fang var.Pingguoensis and C.pingguoensis D.Fang var.terminalis,which includes almost all populations;the total length of chloroplast DNA fragments?cpDNA?is 5190 bp,and seventeen polymorphic loci are detected,resulting in a total of nine haplotypes,C.pingguoensis D.Fang var.Pingguoensis and C.pingguoensis D.Fang var.Terminalis have no shared haplotypes based on cpDNA.2)Based on microsatellite markers,the results showed that there was a high degree of genetic differentiation among the populations of C.pingguoensis,and the genetic differentiation coefficient(F_ST)between populations was between 0.1331 and 0.6337,Among them,F_STT of only one group was less than 0.15;the differentiation coefficient(F_ST)of ten groups was between0.15 and 0.25,and the F_STT value of the remaining forty-four groups was bigger than 0.25.The gene flow between the populations is very low,and gene flow of the four populations is bigger than one,values of gene flow of the remaining fifty-one populations is less than one.The results of STRUCTURE of C.pingguoensis D.Fang var.Pingguoensis showed that there was a high level of differentiation among the populations,and K=7 was the best grouping.At this time,each population was independently grouped.The result of STRUCTURE of C.pingguoensis D.Fang var.terminalis also showed that there was a large differentiation between the populations,K=4 is the best grouping,that is,each population is grouped together.The results based on DNA sequence also showed that the genetic differentiation of C.pingguoensis was very large.The results of AMOVA of the nuclear gene PAL showed that the genetic variation mainly existed between the two varieties,up to 52.74%,13.36%of the variation existed among the populations within the group,the genetic variation within the population was 33.91%,and the genetic differentiation coefficient between populations was0.66094.The results of AMOVA based on cpDNA showed that the variation between the two varieties and the genetic variation between populations were 28.68%and 71.32%,respectively.There was no genetic variation within populations.The differentiation coefficient between the two varieties was 0.28682,and the genetic differentiation coefficient between populations was 1.3)In this study,the results of STRUCTURE showed that K=2 is the best genetic grouping.At this time,the two varieties are grouped solely;the network diagram of the single copy nuclear gene PAL and cpDNA is roughly divided into two groups,C.pingguoensis D.Fang var.Pingguoensis and C.pingguoensis D.Fang var.Terminalis were integrated into each group;The above results indicated that the genetic differentiation between the two varieties was very high.4)The results show that populations of C.pingguoensis D.Fang var.Pingguoensis and C.pingguoensis D.Fang var.Terminalis are divided into two groups,It is recommended to divide these populations into two management units for protection.populations of C.pingguoensis D.Fang var.Pingguoensis and C.pingguoensis D.Fang var.Terminalis have great differentiation,Therefore,when in situ conservation was carried out,multiple populations should be protected as much as possible;if lt's ex situ conservation,the number of samples of every population should be increased as much as possible.