烟草单染色体的分离与扩增及其SSR的检测

Isolation and amplification of single chromosomes and detection of their SSRs in tobacco

以普通烟草根尖为材料,在显微操作仪下分离单染色体,并用LAM-PCR方法进行扩增.对扩增产物用点杂交进行鉴定,用Southern杂交确定目的扩增片段的大小,并用本实验室开发的SSR特异引物检测SSR位点.结果表明:获得的单染色体扩增产物均存在来自烟草基因组的序列,扩增产物大小为250-3000 bp;利用SSR特异引物可以从单染色体扩增产物中检测到目标SSR位点.

With root tips of tobacco（Nicotana tobacum L.） as material,single chromosomes were isolated under the microscope operating instrument and amplified using LAM-PCR（linker adaptor-mediated PCR） method.The amplified products were examined by dot hybridization and their sizes were determined by Southern hybridization.Meanwhile,SSRs in the amplified fragments were detected with specific SSR primers developed by ourselves.The results showed that the amplified products of single chromosomes contained sequences from the tobacco genome and the sizes of amplified products approximately ranged from 250 to 3000 bp;SSRs could be detected from the amplified products of single chromosomes using the specific SSR primers.The results of this study will facilitate further application of the isolation and amplification technology in tobacco.