Genetic Characteristics Of SNP In Genes Related To Rubber Biosynthesis And Latex Flow And HbREF3 Expression Analyses In Hevea Germplasms

Natural rubber(NR)is mainly produced from rubber tree(Hevea brasiliensis).The low self-sufficiency rate is one of the main problems in China’s NR industry.As a typical resource-constrained industry,irreplaceable in planting area and product performance.In China,the planting area of rubber tree is limited and almost utilized,therefore,increasing latex yield per unit area is the only way to improve the natural rubber self-sufficiency rate of NR.Breeding rubber tree varieties with high-yield and high resistance to stresses is an effective way to increase latex yield.At present,elite varieties of rubber tree are mainly cultivated by traditional breeding method in China,and the elite varieties have gone through the selection and breeding process from primary variety to secondary one or more than secondary one.Traditional breeding method of rubber tree is time-consuming and low efficiency,whereas evaluation of rubber tree germplasms,the genetic composition of rubber tree varieties,the genetic characteristics of parents and offsprings,the gene expression patterns,etc are prerequisite and foundation for improving breeding efficiency,which can provide theoretical guidance for the efficient implementation of rubber tree breeding.In this study,the genes related to rubber biosynthesis and latex flow were selected as research objects,and the core parents(RRIM 600 and PR 107)and 9 offsprings of the core parents,4 other Hevea species and 9 wild species were selected as research materials.Transcriptome sequencing data of genes related to rubber biosynthesis and latex flow were analyzed between the parents and their offsprings.After elucidating characteristics and inherited features of SNPs in core parents,we analyzed and validated genetic makeup and characteristics of SNPs between parents and their offsprings.Some SNPs were selected to analyze genetic makeup among other Hevea species,wild species and cultivated species.In addition,expression patterns of HbREF3 were compared with in other Hevea species,wild species and cultivated species.The main results are shown as following.In total,259 SNPs in 31 genes related to rubber biosynthesis and latex flow were identified in RRIM 600 and PR 107 by analyzing transcriptome data from two parents and 9 offsprings.Among the 259 SNPs,the numbers of SNP transitions and transversions were 151 and 108,accounting for 58.3%and 41.7%,respectively.SNPs of parentsmutation type,the numbers of SNP transition types,T?G and G?C.was 73 and 78,accounting for 28.19%and 30.12%respectively;the numbers of SNP transversion types,C?A,T?G,G?C,and T?A,were 31,20,16,and 41,accounting for 11.97%,7.72%,6.18%,and 15.83%.respectively.363 SNPs identified with transcriptional sequencing data were selected to verify by sequencing the products of PCR amplification between parents and their offsprings.The results showed that 330 SNPs were consistent with the results from transcriptome sequencing data,accounting for 92.56%of SNPs accurancy.SNP consistencies of core parents and their offsprings were 98.48%and 91.25%respectively.The results mentioned above suggested that SNPs identified by transcriptome sequencing data were highly reliable in this study.Characteristics and inherited features of SNPs in genes related to rubber biosynthesis and latex flow between parents and their offsprings were as followings:SNP heterozygous percentages of PR 107 and RRIM 600 were 77.61%and 73.75%,respectively;SNP heterozygous percentages of the nine offsprings were between 60.62%and 82.63%.Heterozygous percentages of 4 offsprings,RY 7-33-97,BT 155,BT 3410,and DF 7825,were higher than ones of their parents,and the rest of the offsprings,RY 7-20-59,DL 2138,RY 618,RY 6231,and WC 11,were lower than ones of their parents.SNP genotypes of the genes related to rubber biosynthesis and latex flow showed that the SNP genotypes of RY 7-20-59 were more similar to PR 107;BT 155,BT 3410,DL 2138,and RY 6231,were more similar to RRIM 600;the rest of the offsprings,RY 7-33-97,RY 618,DF 7825,and WC 11,were equally similar to those of parents PR 107 and RRIM 600.The genetic makeups of 33 SNPs in genes related to rubber biosynthesis and latex flow among 4 other Hevea species,9 wild species and 11 cultivated species indicated that SNP heterozygous percentages of other Hevea species largely changed,from 57.58%to 87.88%;the heterozygous SNPs percentage of 9 wild species was between 63.64%and 72.73%;among the 11 cultivated species,SNP heterozygous percentages of primary and secondary clones were separately 60.47%and 74.42%,and tertiary clones were between 87.88%and 96.97%.Generally speaking,SNP heterozygous percentages of tertiary clones were higher than those of 9 wild species and 4 other Hevea species except Hevea benthamiana.Compared with cultivated species,the average PIC of wild species and other Hevea species were lower and higher than that of the cultivated species,respectively.The expression levels of HbREF3 in 4 other Hevea species,9 wild species,and 11 cultivated species were as following:among 11 cultivated species,expression levels of HbREF3 in RRIM 600 and PR 107 were approximately equal to each other;HbREF3 expressions in WC 11 and BT 3410 were higher than those of their parents;HbREF3 expression in RY 618 was almost equal to its parents PR 107;expression levels of HbREF3 in DF 7825,DL 2138,Y 7-20-59,RY 7-33-97,RY 6231,and BT 155 were lower than those of their parents RRIM 600.Expression levels of HbREF3 in 4 other Hevea species were lower than those of PR 107 and RRIM 600.Among 9 wild species,HbREF3 expression in RO/J/5 was slightly higher than that in PR 107,and the expression levels of HbREF3 in the rest of wild species were lower than RRIM 600.