Study On The Germplasm Innovation For Grape Breeding Seedlessness With Rose Aroma Using Embryo Rescue

Seedless grapes have become the first choice for consumers to choose fresh fruits and preserved fruits because of their sweet taste and ready-to-eat.Aroma is an important factor affecting fruit quality.At present,there are still very few seedless and fragrant grape varieties that can be widely promoted and cultivated in the market.Therefore,a new variety that combines the two traits of seedless and aroma is an important goal in grape breeding research.The use of embryo rescue technology to select seedless grape varieties as the female parent and equip the grape varieties with a strong rose fragrance as the male parent.Obtaining stable genetic progeny lines after crossing is of great significance in the cultivation of high-quality fruit tree germplasm resources.In this study,the expression levels of key enzymes for terpene synthesis were analyzed by fluorescence real-time quantitative PCR,and 5 rose fragrance varieties were identified as hybrid male parents,and 9 seedless grape varieties were selected as hybrid female parents,and a total of 12 seedless roses were configured.Fragrance hybrid combination for embryo rescue in vitro culture.The use of seedless molecular markers for early-assisted selection and identification of seedless traits in hybrid progeny.The main findings are as follows:1.Selection of fragrance parents:The unique smell of Muscat grape varieties is related to the presence of terpenoids.Using the ripe fruits of 8 table grapes including Muscat Hamburg and 183 as the materials.Detect the expression of key enzyme genes in terpenoid synthesis pathway in grape berries with different flavors.The key enzyme genes DXS1,DXS3,DXR and HDR,which regulate the early stages of the synthesis pathway of terpenoids,are expressed in the highest expression levels in’Shine-Muscat’,’183’and’Muscat Hamburg’respectively.The key enzyme genes GPPS and FPPS,which control the mid-term of the terpenoid synthesis pathway,have the highest expression levels in’183’and’Shine-Muscat’respectively.The key enzyme genes Linnalool syn andα-Terpineol syn,which regulate the late stage of the terpenoid synthesis pathway,have the highest expression levels in’Shine-Muscat’and’183’respectively.Compared with other enzyme genes,the expression level ofα-Terpineol syn in each mature grape fruit is also the highest,which is 10-65 times the expression level of other enzyme genes.2.Configure seedless fragrance hybrid combination:According to the gene expression results of key enzymes for terpene synthesis,the varieties with higher expression levels of terpene synthase genes were screened out,and 12 hybrid combinations were configured.Using embryo rescue technology,477 hybrid lines were obtained,and 327 survived.The scented seedless embryo rescue hybrid combination with’Ruby seedless’and‘Fresno Seedless’as the female parent has a better embryo saving effect.The scented seedless embryo rescue hybrid combination with‘Muscat Hamburg’and‘183’as the male parent had a higher seedling rate.The embryo development rate and seedling rate of Ruby‘Seedless×Shine-Muscat’are relatively highest,which are 55.63%and 38.82%respectively.‘Ruby Seedless’and‘Fresno Seedless’are suitable as female parent materials for embryo rescue breeding,and‘Muscat Hamburg’and‘183’are suitable as male parent materials.3.The best sampling time is determined:Continuous sampling of the configured seedless fragrance embryo rescue hybrid combination.The hybrid combination with’Jingzaojing’as the female parent had a higher embryo development rate at 36 and 42days after pollination,and the highest rate of embryo development reached 13.33%at 42days after pollination.The hybrid combination with’Fresno Seedless’as the female parent had the highest embryo development rate and seedling rate at 36 days after pollination,which were 31.25%and 33.33%,respectively.The hybrid combination with‘Australian seedless’as the female parent had the highest embryo development rate at 37 days after pollination,reaching 10.42%.4.Optimization of embryo germination medium composition:An optimized embryo germination medium:WPM+0.1 mg·L^-1 BR+0.2 mg·L^-16-BA.The germination rate was the highest when using this medium,55.88%,which was higher than the control 21.59%.Adding two exogenous hormones,BR and IAA at the same time,can inhibit embryo germination.The root growth of hybrid embryos with BR added was stronger than that of hybrid seedlings without BR.5.Use the seedless gene probe GSLP1-569 and the seedless molecular marker SCF27-2000 to identify 56 F₁ lines for early seedless traits,and 14 of them carry the specific bands of the above two types of seedless molecular markers.It is preliminarily judged that these 14 hybrid strains carry seedless traits.In summary,this study obtained a total of 327 seedless rose fragrance hybrid strains,56of which were identified for early seedless traits,and 14 strains simultaneously carried two specific bands with different seedless molecular markers;Two dominant female parent varieties,’Ruby Seedless’and’Fresno Seedless’,are produced.Combined with the expression results of the key enzyme genes in the synthesis of terpene in the parent,the scented embryos that are suitable for use as the male parent for the rescue breeding of scented embryos are‘Muscat Hamburg’and‘183’.It was determined that the best sampling times for hybrid combinations with‘Jingzaojing’,‘Fresno Seedless’and‘Australian seedless’as the female parent were 42 days,36 days and 37 days after pollination.An optimized embryo germination medium:WPM+0.1 mg·L^-1 BR+0.2 mg·L^-1 6-BA.