Genetic Diversity Of Bashania Fangiana Clonal Populations With Two Different Genet Ages

Bashania fangiana is a perennial monocarpic species, which has both types of sympodial and monopodial rhizome systems in a genet. As a typical clonal plant, B. fangiana flowers once about 45-55 years or even more seldom. After mass death, its populations recover mainly by the development of seedling cohorts. When the seedlings reach full size stage, they extend rhizomes and produce culms, resulting in dense populations again. B. fangiana is one of the staple food bamboos for the giant panda (Ailuropoda melanoleuca), which is crucial to maintain its survival and reproduction. Amplified fragment length polymorphism (AFLP) fingerprints were used to reveal genetic diversity, clonal structure and the relationship with succession stage of B. fangiana clonal populations with two different genet ages (Ni,≤30 years versus N2,≥70 years) at Wolong National Natural Reserve, which can also discover the numbers, size and spatial pattern of clone in populations, so provide the basic data for the further research of B. fangiana in clonal growth, distribution characteristics and ecological adaptation, and provide some reference materials for the study of giant panda, such as staple food bamboo and population protection.We obtained AFLP binary matrices from 96 B. fangiana ramets samples. The observed numbers of bands by AFLP analysis were 202 using the ten selective primer pairs. A total of 111 polymorphic bands were identified from the both populations, and each primer pair produced an average of 11.1 polymorphic bands. The average proportion of polymorphic loci was 53.9%. At the population level, the polymorphic bands in Ni population was 107, and the average proportion of polymorphic loci in Ni population was 51.87%. The corresponding values for the N2 population were 91 and 44.33%. In the proportion of polymorphic loci view, genetic diversity in Ni population was slightly higher than that in N2 population.The use of POPGENE3.2 software analysis showed that there were high genetic diversity in B. fangiana clonal populations, and the percentage of polymorphic loci (PPB), observed number of alleles (na), effective number of alleles (ne), Nei’s gene diversity (h) and Shannon’s information index (Ⅰ) were 54.95%,1.5495,1.2239,0.1428 and 0.2265, respectively. At the population level, the values of PPB in two B. fangiana clonal populations were 52.97%(N1) and 45.05%(N2), respectively. The other four indices (na, ne, h and Ⅰ) in N1 population were all higher than that in N2 population, but the difference was not significant. The coefficient of gene differentiation (Gst) among the two populations was 0.0571, and the estimate of gene flow (Nm) was 8.2639. This results demonstrated that genetic variation within populations was larger than that among populations, and gene flow among the populations was big. So, the genetic differentiation among the populations was not obvious. Our analysis of molecular variance (AMOVA) consistently showed that a large proportion of the genetic variation (87.79%) resides among the individuals within populations, whereas only 12.21% are found among populations.The results of genetic distance (D) and genetic identity (Ⅰ) discovered that the genetic basis between the two B. fangiana clonal populations was relatively similar. The genetic identity was up to 0.9813, while the genetic distance was only 0.0189. The UPGMA cluster analysis revealed two distinct groups:Ⅰ, the 41 ramets sampled from population Ni; Ⅱ, the 48 ramets (number from 49 to 96) sampled from population N2. Ramets of the same B. fangiana population first clustered into one group, then clustered with other population. Only a small number of ramets (ramets 19,26,27,28,32,33 and 37) did not gathered together with other ramets in the same population (N1). Results of the UPGMA analysis also revealed that the high genetic similarity were shared between the two populations, which was consistent with the results of genetic identity. In PCA based on the AFLP data matrix of the 202 bands for 96 ramets in two B. fangiana populations, the first three components effectively distinguished the two populations and ramets of the same population gathered close,.which was consistent with the results of UPGMA.Samples showing the same multilocus AFLP phenotype were considered to be the same clone (or the same genotype). Among 96 B. fangiana samples,92 genotypes (G=92) with identical AFLP fingerprints were detected. The number of genets in N1 and N2 populations was 47 and 45 respectively. Our results demonstrated that clonal diversity in B. fangiana populations were high, the mean proportion of distinguishable genotypes (G/N) and Simpson’s index of diversity (D) for B. fangiana was 0.9583 (N1=0.9792, N2=0.9375) and 0.9982 (N1=0.9991, N2=0.9973), respectively. Estimates of genotypic evenness (E) in two populations were 0 (N1) and 0.6667 (N2) respectively. Our results showed that both of the sampled populations were multiclonal, and the genotypic diversity and genet density of B. fangiana clonal population did not change significantly with genet aging. Results of the clonal structure analysis revealed that B. fangiana populations are not dominated by one clone or have any dominate genotype and we did not observe any mixed distribution of multiple clones. The largest size of a single clone might occurs a distance of about 30 m at least. No common genotypes were found among the individuals from the two populations, showing a great clonal differentiation between them.In conclusion, the clonal diversity of B. fangiana population was reasonably high, contrary to the general expectation B. fangiana clonal populations have higher genetic diversity which were maintained through time. In addition, the high level of genotypic diversity in the two populations implies that the further works were needed to investigate the reasons for the poor seed set in B. fangiana after flowering.