2020-04-012024-05-14https://scholars.lib.ntu.edu.tw/handle/123456789/660195摘要：栽培作物的病蟲害抗性基因或對非生物性逆境的耐受性基因通常來自野生物種。但在野生親緣種中與目標性狀緊密相連的不佳性狀基因所引起的連鎖累贅，對所需性狀的滲入可能會造成阻礙。尤其當滲入的抗性基因與重要園藝性狀緊密相連，且基因座的重組頻率低時，即使重複進行回交也無法使表型恢復。世界蔬菜中心已培育出栽培種番茄和抗蟲野生親緣種(S. galapagense)之間的種間雜交BC3F3族群，該抗蟲BC3F3品系具有很小的果實，且通過與栽培種番茄反復回交並無法成功提高果實大小。由於昆蟲抗性基因座位於2號染色體上且與果實大小決定基因fw2.2緊密相連，此漸滲系可能攜帶fw2.2的野生等位基因而導致果實較小。本計畫將利用CRISPR / Cas9技術，誘導fw2.2基因編碼區和調控序列產生突變，使fw2.2基因座恢復成栽培種的等位基因。突變植株將被再生並進行外表型分析，以比較植株生長、形態特徵以及果實大小的變化。藉由透徹的序列和表達分析，將能得知更多與調控番茄果實大小相關的資訊，除了能在fw2.2基因上選擇更適合的突變目標，亦能尋找其他潛在的突變目標，甚而找出由CRISPR / Cas9基因編輯技術所導致的非目標突變。<br> Abstract: Resistance genes against pests and diseases or tolerance genes for abiotic stresses for crop plants are often sourced from wild species. Introgression of the desired trait from wild relatives can be hindered by linkage drag caused by unwanted genes tightly linked to the trait of interest. Repeated backcrossing may not effective, if the linkage between the introgressed resistance gene and an important trait of the cultivated parent is too tight and recombination frequency at the locus is low. The World Vegetable Center has generated a backcross population from an interspecific hybrid between cultivated tomato (Solanum lycopericum) and an insect resistant wild relative (S. galapagense). The resultant insect resistant backcross lines have very small fruits and improving the fruit size through backcrossing was not successful. The insect resistance trait introgressed into chromosome 2 of this line is tightly linked to the fruit size gene fw2.2, consequently, insect resistant backcross lines carry the wild allele of fw2.2, causing small fruits. The project proposes to use CRISPR/Cas9 to restore the cultivated allele at the fw2.2 locus by mutating coding and regulative sequences of the fw2.2 gene. Mutant plants will be regenerated and phenotyped for developmental and morphological traits and for variation of fruit size. Thorough sequence and expression analysis will optimize the selection of the target for mutation in fw2.2, provide information on potential further mutation targets for manipulating fruit size trait in tomato and pinpoint possible non-target mutations introduced by CRISPR/Cas9 gene editing.番茄基因編輯果實大小種間雜交種tomatoCRISPR/Cas9 gene editingfruit sizeinterspecific hybrid