Study Of Molecular Markers On Important Quality Traits In Sea Buckthorn Germplasm

The quality traits are key factors in breeding of sea buckthorn.In recent years,RNA-Sequencing technique has been developed and applied in woods.New type of DNA markers such as SSR markers were developed more and more,which give the method basis to the association analysis of important quality traits.This study used eighty sea buckthorn cultivas/lines as materials,which were from different genetic backgrounds.The important quality traits were analyzed,such as hundred-grain weight of berries and seeds,oil content in seeds and pulp,fatty acid composition in seed oil and pulp oil,vitaimin C content,the composition and content of vitamin E and carotenoids.At the same time,simple sequence repeats（SSR）primers for sea buckthorn were developed by RNA-Sequencing technique,which were used for genetic relationship analysis among sea buckthorn germplasm and association analysis with fruit quality traits.Furthurmore,this study used twenty-two sea buckthorn accessions with different disease indices to deied-shrink disease（DSD）as materials,and performed the genetic relationship analysis with ISSR and SRAP markers.Then,stepwise MRA method was applied to seek markers associated with DSD.1.There are significant differences in quality traits among eighty sea buckthorn cultivars/lines from different genetic backgrounds.The hundred berry weight（HBW）and hundred seed weight（HSW）of samples of ssp.mongolica were biggest at the average value of 47.38±10.72 g%and 1.60±0.28 g%,respectively.The berry shape indice（BSI）of three groups were also significantly different（p<0.001）,oblong for ssp.mongolica,oblate for ssp.sinensis and circular for hybrid varieties.The berries of ssp.sinensis with small size contained the highest vitamin C in juice and those of ssp.mongolica with bigger size had low content of vitamin C in juice.The proportion of linoleic andα-linolenic acids were high in seed oil at 30-40%and palmitic and palmitoleic acids were dominant fatty acids in pulp oil at 20-40%.Vitamin E in sea buckthorn berries mainly composed of four tocopherol isomers andα-tocopherol was the major isomer in pulp oil and seed oil.The carotenoids composition was complicated in matured fruits.β-Carotene were the main component in sea buckthorn berries.2.According to the results of RNA-Sequencing with mixed samples of roots,stems,leaves,and fruits,SSR markers primer pairs were designed and fourty SSR markers with polymorphism were developed which were suitable to sea buckthorn.We firstly applied these SSR markers to analyze the relationships among sea buckthorn germplasm.The results showed that these SSR markers were suitable for genetic relationship analysis in sea buckthorn germplasm.3.The SSR markers with polymorphism were used to analyze the population structure among eighty sea buckthorn cultivars/lines and perform the association analysis with important fruit quality traits based on GLM and MLM.Nine SSR markers were estimated to assocate with the fruit quality traits.One marker associated with HBW（Unigene14574＿HR＿2027）and one associated with HSW（CL3558.Contig1＿HR＿936）specially.The other seven markers associated with oil content of seeds and pulp（CL2024.Contig1＿HR＿553 and CL1076.Contig2＿HR＿301）,α-linolenic acid in seed oil（CL4716.Contig3＿HR＿1126）,palmitic acid in pulp oil（CL2339.Contig9＿HR＿634）,α-tocopherol in pulp（CL3904.Contig3＿HR＿1006）,and vitamin C content（CL2442.Contig1＿HR＿672 and CL3541.Contig1＿HR＿929）.No markers associated with berry shape indice,linoleic acid andα-tocopherol in seeds,andβ-carotene in pulp were found.4.The DSD largely affects the growth of sea buckthorn and the resistence to it is an important measure of quality.Use ISSR and SRAP analysis and stepwise multiple regression analysis（MRA）method,four ISSR markers(887₁₉₀,835₇₀₀,809₂₉₀,and811280)and eleven SRAP markers(Me1-Em3₆₀₀,Me1-Em1₆₈₀,Me2-Em1₆₅₀,Me2-Em1₉₅₀,Me3-Em6₁₃₀₀,Me2-Em6₃₂₀,Me2-Em6₄₀₀,Me1-Em2₆₀₀,Me1-Em1₁₂₀₀,Me1-Em1₁₇₀₀,Me2-Em2₂₅₀)were significantly found correlated with DSD resistance（p<0.001）.These markers provide an early selection for breeding programs that select lineages with DSD resistance,especially when no other genetic information such as linkage maps and quantitative trait loci are available.To sum up,Hippophae rhamnoides ssp.mongolica cultivars/lines own more economic values by the analysis of quality traits.Fourty SSR primer pairs were screened and applied to analyze genetic relationship among sea buckthorn germplasm using RNA-Sequencing.Several markers associated with resistence to DSD were developed based on ISSR and SRAP with MRA method.These results have important theoretical value and practical significance in the identification of variety.This research will give references to QTL identification of important quality traits and germplasm evaluation with potential economic value.Also the results will lay the theoretical foundation for seeking genes associated quality traits and selection of optimal germplasm in sea buckthorn.