Genetic Diversity Of Germplasm Resources Of Michelia Macclurei Dandy

Michelia macclurei Dandy(Magnoliaceae),also known as Zui Xiang Han Xiao in Chinese,is a large evergreen tree and it is an important local precious timber species with multi-functional benefits in southern China.In view of the lack of information on the genetic diversity and improved variety in M.macclurei,15 quantitative traits and 4 qualitative traits were systematically assessed using near infrared spectroscopy,transcriptome sequencing technologies and EST-SSR molecular markers from the phenotype,physiological and biochemical and molecular levels in order to characterize genetic diversity in various aspects among germplasm resources and analyze the genetic variation at the provenance and family levels.These results can provide a theoretical basis for selection of outstanding genetic materials and establishment of breeding program in order to promote the planting in the tropical and subtropical regions.The main results from this research are summarized as follows:(1)There were highly significant differences in growth traits,density of permanent branch(DPB)and thickness of permanent branch(TPB)among provenances and among families(P<0.01)indicating great potential for genetic improvement at provenance and family levels.However,stem form straightness(SFS)and stem form axis persistence(SFA)were not significantly different among provenances or families.The average genetic diversity index of growth traits was 2.038 indicting high genetic diversity,while that for form traits was only 0.805 indicating lower level of genetic diversity.Based on the individual heritability,SFS,SFA and DPB were almost not controlled by genetic;TPB was under low genetic control;tree height(H),diameter at breast height(DBH)and volume(V)were under moderate genetic control;and crown width(CW)was under high genetic control.Based on the family heritability,H,DBH,V,CW and TPB were all under high genetic control.With 20% of the selection rate,the genetic gain of main traits was 3.14~6.23% for H,7.37~18.78% for DBH,10.14~33.01% for V,8.73~21.81% for CW,and 3.82~12.02% for TPB.Genetic correlation and phenotypic correlation in H,DBH,V and CW were highly and positively significant.Significant negative correlations were observed between latitude and H,DBH,V and CW,however,it was significantly positively correlated with TPB.There was a significant positive correlation between altitude and DPB.(2)Wood basic density(BD),fiber width(FW)and fiber length-width ratio(LTW)were highly significantly different among provenances and among families(P<0.01).Fiber length(FL)was also highly significantly different among families(P<0.01)but not among provenances,reflecting a strong potential for selection and breeding.The genetic diversity index of each wood property averaged 2.064,indicating high genetic diversity.High individual heritability and family heritability values in BD,FL and LTW suggested these traits were under high genetic control.The individual heritability and family heritability in FW was lower thus under moderate genetic control.With 20% of the selection rate,the genetic gain of each wood properties was 2.43~5.12% for BD,4.02~9.60% for FL,1.03~2.61% for FW and 2.73~6.86% for LTW.Correlation analysis showed that longitude was significantly and positively correlated with FL and LTW.There was a significant positive correlation between BD and all growth traits,but there was a significant negative correlation between FW and all growth traits,and a significant negative correlation between FL and FW.The BD of M.macclurei could be selected jointly with growth traits,while FL should be independently selected.(3)The prediction model of nitrogen(N),phosphorous(P)and potassium(K)content of leaves using the near infrared spectroscopy were better,which can be used to predict the mineral nutrition in the leaves of M.macclurei,and the prediction model of N was the best.The N and K contents in the leaves were significantly different among provenances and among families.There was significant difference in P among families,but not among provenances.There were significant differences in leaf length(LL),leaf width(LW),length-width ratio(L/W)and leaf area(LA)among provenances and among families(P<0.01).The genetic diversity index of leaf traits was high with an overall average of 2.082,showing higher genetic diversity.The LL,LW and LA of M.macclurei were significantly positively correlated with H,indicating that these leaf traits could be used as auxiliary selection index for the growth rate of M.macclurei.N,P and K were significantly and positively correlated with elements,and K was significantly positively correlated with LL,LW and LA,and K also was significantly positively correlation with H.N,and P were positively correlated with longitude.There were significant positive correlations between latitude and LL,LA,N and P.There was a significant positive correlation between elevation and LA.(4)Using the multi-objective decision method,comprehensive evaluation was performed on the germplasm resources of M.macclurei using H,DBH,CW,TPB,LA,K,BD and LTW.Provenance 13,provenance 2,provenance 3 and provenance 4(total 4 provenances)were superior provenances.Family 10,family 36,family 78,family 17,family 16,family 45,family 2,family 82,family 52,family 25,family 15 and family 61(total 12 families)were the better families.When selecting the superior families,the selection rate was 12.24%.The average genetic gain of the main traits was 3.50% for H,7.37% for DBH,6.45% for CW,1.49% for BD and 2.04% for LTW.From comprehensive assessment,provenances from Guangdong Province were superior to provenances from Guangxi,and the two provenances from Gaozhou performed well;the provenances from Yulin,Guangxi Province were poor.However,at the family level,some families from Guangxi also performed better.(5)In order to develop and analyse SSR makers of M.macclurei,transcriptome sequencing was performed on the roots,stems,and leaves of M.macclurei and 97 503 Unigenes were obtained.The average length was 852 bp and N50 was 1 010 bp.The sequences obtained were of relatively higher quality and contained a large amount of information.A total of 61 205(62.77%)Unigenes were successfully annotated to COG,GO,KEGG,Swiss-prot and NR databases.The differentially expressed genes analysis showed considerable differences between the roots,stem and leaves,and less differentially expressed genes between the stem and leaves.Through the exploration and analysis of SSR loci in Unigenes,21 218 SSR containing sequences were found and 27 379 SSRs were obtained,with a frequency of 28.08% and an average of about 1 SSR per 3 kb.Mononucleotide and dinucleotide repeat were the main SSR types of M.macclurei.Among all the 85 repeat motifs,the highest proportion of(A/T)n was 41.65%.The variation of the length of mononucleotide repeat SSR was the most abundant,followed by the dinucleotide.The SSR loci in the M.macclurei transcriptome showed higher frequency and density of distribution,rich repeat motifs,higher repeat times,more long fragment and significant potential of polymorphism.The 12 EST-SSR markers were applied to study of genetic polymorphism for 10 natural populations of M.macclurei.A total of 162 alleles were detected,averaging at 13.5 alleles per locus with an average effective allele of 6.161;The average Shannon diversity index(I)was 2.001,and the average polymorphism information(PIC)per locus was 0.802,which locus polymorphism was higher level;the population diversity was also higher,and the average expected heterozygosity of populations(He)was 0.763.The average of Shannon diversity index(I)was 1.725.The level of differentiation among populations was lower,the average differentiation coefficient(FST)of the 12 SSR loci was only 0.061,the average of gene flow(Nm)was 4.384,and the percentage of variance components of the populations in the Analysis of Molecular Variance(AMOVA)accounted for only 3.656%,indicating that the majority of genetic variation was within population.The average genetic identity(Gi)of the populations was 0.813,and the average genetic distance(Gd)was 0.214.There was no significant correlation between geographic distance and genetic distance among populations;UPGMA clustering of M.macclurei populations by genetic distance could be divided into two major groups and 4 subgroups.