利用基因組原位雜交和異源染色體添加系建立Nicotiana sylvestris的核型

Abstract

Nicotiana sylvestris為栽培菸草的母本，共有十二對染色體，多為中位中節和近中位中節；其染色體長度和長短臂比例相似，且染色體上缺乏明顯的細胞遺傳標記，所以一直無法建立正確的核型。本實驗以N. sylvestris基因組DNA為探針，N. plumbaginifolia基因組DNA為覆蓋DNA，與N. sylvestris根尖細胞進行基因組螢光原位雜交(genomic in situ hybridization, GISH)；結果顯示雜交訊號在N. sylvestris染色體的分佈有強弱的差異，且染色體上訊號的多寡和位置亦不同。根據基因組原位雜交結果，以及染色體長度和長短臂比例、衛星體的有無，並輔以本實驗室建立的N. plumbaginifolia-sylvestris異源單染色體添加系，我們建立一個正確的N. sylvestris的核型，未來能應用於栽培菸草的基因組演化的分析，添加系中變異型N. sylvestris 染色體的分析，DNA序列的實質定位，以及N. sylvestris中串聯性重複序列的定位等研究上。

Nicotiana sylvestris (2n=24) is the maternal parent of cultivated tobacco (N. tabacum). The lengths and arm ratios of N. sylvestris chromosomes are similar, and there are no obvious cytological markers on the chromosomes. Therefore, it is difficult to identify the individual chromosomes by traditional cytogenetic techniques. In this study, we hybridized somatic metaphase chromosomes of N. sylvestris and N. plumbaginifolia-sylvestris monosomic addition lines using genomic DNA of N. sylvestris as a probe and unlabeled genomic DNA of N. plumbaginifolia as a block. The results show that patterns of hybridization signals in individual N. sylvestris chromosomes are distinctly different. Based on patterns of genomic in situ hybridization (GISH), chromosome lengths, arm rations, and positions and sizes of satellites, we construct a karyotype for N. sylvestris. The karyotype is useful for investigation of the evolution of cultivated tobacco, identification of chromosomes in aberrant addition lines, and localization of repetitive, single and low-copy sequences.