Construction Of Wild Cucumber Substitution

Cucumber production contributes significantly to vegetable supply in China. As the scale of protected cultivation grew in past 30 years, loss to disease and insect increased accordingly, which became the main constraints to the yield and quality for cucumber. Gain of breeding is limited using the current parental lines due to the narrow genetic base of cucumber, lack of resistance in parental lines, and the time-consumption of the traditional breeding approaches. Introgression of wild alleles into elite inbred lines is a must.In late nineteenth century, British botanist J. D. Hooker discovered wild cucumbers in North India, Sikkim and other places around of the Southern Himalayas area. He named it subsequently as Cucumis hardwickii. Compared with the cultivated cucumber, the wild cucumber has a unique morphology, including small leaves, no significant main vine, highly branched, more fruit setting, small and oval fruit, bitter taste thus not edible, sparse and black spikes, but good resistance to disease and stress C. hardwickii and C. sativus can cross and their offsprings are fertile. Therefore, it is generally agreed that the wild cucumber is a cucumber subspecies (C. s. var. hardwickii). The wild cucumber may be wild ancestor of the cultivated cucumber (C. s. var. sativus).Chromosomal segment substitution lines, (CSSL) are a series of near-isogenic lines that carry single fragment from the donor genome and jointly cover the entire donor genome. CSSL population can be constructed via marker-assisted backcrossing and selfing. Except for a substituted segment from the donor parent, the other parts of the genome in these lines are the same as the recipient parent. Therefore, any phenotype differences between the substitution lines and the recipient parent can attribute to the substitution fragments. Studying of quantitative traits is simplified using CSSL as individual QTL can be viewed as single Mendelian factors.So far, construction of CSSL population in cucumber has not been reported. Cucumber genome sequencing paves the way to build high-density molecular marker linkage map of cucumber and subsequently, to construct CSSL population. In this study, we used the C. s. var hardwickii line PI183967 as the donor material, the elite C. s. var sativus inbred Xintaimici as the recepient material, to generate the first CSSL population of cucumber.Based on the high-density genetic map of cucumber, we chose the 140 pairs of SSR markers which uniformly distributed in the seven chromosomes, then detected polymorphism between the wild line PI183967 and the inbred Xintaimici. This resulted in 62 polymorphic SSR markers between parents. The rate of marker polymorphism is 37.1%.The 62 SSRs were used to scan the genome of 79 BC1 plants. Genotype display software GGT were used to visualize the genomic constituents of the BC1 population. On average the wild genomic fragments take up the 47.3% of the genome of one BC1 plant. We selected 18 lines from the 79 lines, which provides a coverage of the wild cucumber genome for about four times. The 18 plants were used for making the BC2 generation. All the 352 BC2 plants were analyzed with the 62 SSRs. On average the wild genomic fragments take up the 19.4% of the genome of one BC2 plant. We selected 19 lines to make the BC3 generation, which provide a coverage of the wild cucumber genome for about two times.The BC3 generation consists of 256 individuals, which again were scanned by the SSR markers. On average the wild genomic fragments take up the 7.8% of the genome of one BC3 plant. We selected 31 plants from the 256 lines. Among the 31 plants, 21 carry a single fragment derived from the wild cucumber and 10 carry double wild fragments. In total, the 31 selected plants cover 96.81% of wild cucumber genome. These plants were selfcrossed to generate the BC3S1 population that allow single wild fragment to become homozygous, enabling further phenotyping and genotyping studies.The CSSL population provides a new resource for use of the advantageous wild cucumber genes, enables an accurate analysis of quantitative traits, and also contributes to the study of molecular mechanism underlying the domestication process of cucumber.