基于染色体涂染的甜瓜属主要物种染色体结构分化研究

Cucumis genus includes two economically important crops, cucumber (C. sativus L.) and melon (C. melo L.), and other wild species as well, such as C. metuliferus L., and C. hystrix Chakr. which carry important resistant genes. The utilization of useful genes from wild species for crop improvement has been hampered due to the extensive cross-incompatible among most of cucumis species, or the severe recombination suppression between parental chromosomes in the interspecific hybrid. The structural differentiation of chromosome among species could explain these obstacles. This research aims at elucidating the characterization of structure differentiation of chromosomes in cucumis, which could provide theoretical guidance for the introgression breeding. Comparative chromosome painting allows the direct visualization of chromosome structure differentiation, like chromosome rearrangements, duplication/deletion, etc. In this research, single copy gene chromosome painting technology (ScgCP) will be employed to investigate the structure differentiation of chromosome in cucumis. The annotated genes from cucumber and melon genomes will be used for designing chromosome-specific probes. Comparative chromosome painting method will be carried out to detect the homogenous regions among the different species from cucumis. Based on the analysis, the comparative chromosome maps will be constructed, and the structure differentiation of chromosomes of cucumis species will be revealed. The expected results will be helpful for guiding the introduction of useful genes from wild relatives into cultivars based on interspecific hybridization, and provide the basis for elucidating the evolution of Cucurbitaceae species as well.

甜瓜属包括黄瓜和甜瓜两种重要栽培作物，以及非洲角黄瓜、酸黄瓜等多种携带重要抗性基因的近缘种。物种间存在不同程度的杂交不亲和性或杂种染色体组间的重组抑制，阻碍了近缘物种间优异基因的转移利用。染色体结构的差异是造成其生殖隔离或重组障碍的重要因素。本项目研究旨在揭示甜瓜属物种间染色体结构分化特征，为野生资源优异基因的转移利用提供理论指导。物种间染色体涂染的比较分析能直观地反映近缘物种间的染色体重排、区段重复/缺失等结构的差异。项目将基于申请者前期开发的单拷贝基因染色体涂染技术，利用黄瓜和甜瓜全基因组的注释基因，研究特定染色体探针在甜瓜属主要物种染色体上的同源分布，分析物种间染色体的重排特征，构建黄瓜、甜瓜、非洲角黄瓜、酸黄瓜等甜瓜属主要物种间的比较染色体图谱，深入系统分析甜瓜属物种的染色体结构分化特征。预期结果有助于指导甜瓜属种质资源的利用，并为葫芦科作物的进化提供重要理论基础。

本项目利用黄瓜及甜瓜属近缘物种甜瓜、酸黄瓜(Cucumis hystrix Chakr.)、非洲角黄瓜(Cucumis meluliferus)和西印度瓜（Cucumis anguria）为研究材料，通过以荧光原位杂交为基础的染色体涂染方法构建了黄瓜、酸黄瓜、C.anguria以及异源四倍体C. × hytivus的核型。对甜瓜属主要物种染色体重复序列分布特征进行比较，揭示了甜瓜属主要物种重复序列的同源性特征；通过对20个甜瓜属物种的45S rDNA和5S rDNA的比较染色体定位及基因组进化分析，研究基因组结构和物种间染色体进化关系。通过对C. hystrix的亚端粒重复序列CuhySat2和CuhySat3在黄瓜染色体着丝粒位置产生信号特征分析揭示甜瓜属物种染色体的进化机制，有助于理解甜瓜属的染色体进化。开发了单拷贝探针，分析了黄瓜、甜瓜和非洲角黄瓜4号染色体短臂8Mb区段的重排，研究黄瓜7号染色体与C. anguria5号染色体的共线性关系及重排关系。开发了简易的寡核苷酸涂染技术，使用寡核苷酸探针追踪C. × hytivus减数分裂中的染色体，观察种间杂种减数分裂过程中染色体行为，揭示了种间杂种的异源配对情况；通过对寡核苷酸探针进行分段，详细分析了黄瓜-甜瓜-酸黄瓜染色体的重排关系。本项目研究成果对甜瓜属物种染色体及其种间重排关系进行了分析，为彻底了解甜瓜属物种染色体的数量变化打下基础，具有重要的科研意义；同时在研究过程中所开发的单拷贝探针及寡核苷酸涂染技术将成为研究其他植物物种的染色体的重要有力手段。