番木瓜单染色体的显微分离与克隆

Microdissection and Microcloning of Single Chromosome of Papaya

用玻璃针显微分离出番木瓜(CaricapapayaL.)单染色体,经过LA-PCR扩增得到80-700bp的DNA片段。Southern杂交表明,扩增片段与番木瓜基因组DNA之间有同源性,从而证明番木瓜单染色体DNA已经被成功扩增。将扩增产物克隆到pGEM-T-Easy载体中,约获得1.18×105个克隆,酶切鉴定插入片段大小为100-400bp。

Single chromosome of papaya （Carica papaya L.） was isolated by glass needles, and the chromosomal DNA was amplified by LA-PCR （Linker Adapter PCR）. The PCR products were a smear ranged from 80 to 700 bp. Southern blot showed that the PCR products were homogeneous with the papaya genomic DNA, indicating that DNAs from the single chromosome had been successfully amplified. The PCR products were cloned into pGEM-T-Easy vector and about 1.18×10^5 recombinant clones were obtained. The size of the inserted fragments of clones ranged from 100 to 400 bp.