| Flowering, which marks the transition from vegetative to reproductive growth in angiosperms, is dependent on heredity and environmental factors. Under normal conditions, early flowering and reproductive growth can be initiated using less energy than in severe environments, such as low temperatures, and so it is important that flowering plants avoid such conditions in order to survive. Flowering time is a quantitative trait that, in tomato, has high a correlation with fruit ripening and is controlled by a polygene. Since early ripening represents a valuable characteristic for crop production value, there is great interest in elucidating the genetic basis of flowering time. Here, an F2 population generated from crosses between 071-440(late flowering, P2) and Bone MM(early flowering, P1). The major quantitative trait loci(QTLs) related to early flowering were identified using QTL-Seq analysis. The candidate genes were then screened for gene expression, and the expression patterns of two micro RNAs(mi RNAs) were evaluated. The main results obtained are summarized below:1. Seven major QTLs related with early flowering were localized by QTL-Seq and, based on phenotypic investigation of 1,200 F2 plants, DNA bulks from 40 ‘extremely early’ and 40 ‘extremely late’ plants were made. The two progeny bulks and two parent bulks were sequenced, and the QTLs related with flowering time were found to be located between 1.6 Mb and 71.8 Mb on chromosome 1. Based on the single nucleotide polymorphism(SNP)-index, seven major QTLs were predicted: 23.5 Mb-24 Mb, 24 Mb-28 Mb, 35 Mb-40 Mb, 52.5 Mb-54 Mb, 59 Mb-62 Mb, 69 Mb-72 Mb and 70 Mb-74 Mb. Consequently, seven candidate genes were predicted based on the sequence of the tomato genome: EF1(Solyc01g017060), EF2(Solyc01g018000), EF3(Solyc01g034220), EF4(Solyc01g057370), EF5(Solyc01g058640), EF6(Solyc01g073700) and EF7(Solyc01g081360), respectively.2. One QTL was also found using traditional QTL analysis. A total of 143 In Del markers were designed along the QTL region, and 136 F2 plants were analyzed using Map QTL software. The result showed that the QTL was located between Fl-Indel 2 and Fl-Indel 8, which corresponds to a physical region between 23.5 Mb and 25.3 Mb. This region also corresponds to the 23.5 Mb-24 Mb region identified by QTL-Seq; namely EF1.3. PCR primers for the seven candidate genes were designed, and using real-time q PCR it was found that the change in gene expression in parent plants was highest for EF1, with relative expression levels of 219.34(early parent) and 73.46(late parent), respectively. The difference was significant at P￡ 0.05. Thus, EF1, which has sequence similarity to the Ycf2 gene, may be responsible for the early flowering phenotype.4. The expression patterns of both candidate mi RNAs(mi R156 and mi R172) was studied in leaves at different positions(1st, 3rd, 4th, 6th, 9th, and 12 th leaf from the bottom). The expression levels in P1(early) of both mi RNAs increased with consecutive leaf position, and rose substantially in the 12 th leaf, to 1.12 and 4.88 fold, respectively. However, the initial flower of P1 was on 6th leaf. The expression patterns of the two mi RNAs in P2(the initial flower of P1 on 12th) were different. The highest expression of mi R156 was in the 3rd leaf(2.18 fold), while mi R172 expression was highest on the 9th leaf, then decreased in the 12 th leaf to 62.12 and 40.52 fold respectively. It is therefore unlikely that either of the two mi RNAs is involved in early flowering. |
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