CMS高粱保持系叶绿体基因ps1A1和ps1A2片段的克隆与序列比对

CLONING ANG SEQUENCE ANALYSIS OF ps1A1 AND ps1A2 GENES AMPLIFIED SPECIFICALLY FROM THE CHLOROPLAST DNA OF MAINTAINER OF CMS SORGHUM*

以 15种包括同质异核和同核异质高粱材料的总DNA为模板进行RAPD分析 .196个随机引物中 ,引物Y 19扩增得到一个很有规律的差异片段SAY 192 3 0 0 .该片段只出现在 4个保持系 (B)和 3个恢复系 (R)中 ,而不出现在 6个不育系 (A)和 2个杂种F1中 .进一步对cms A1、cms A2两套 8个材料 (A/B/R/F1)的总DNA、mtDNA和cpDNA用引物Y 19进行扩增 ,发现片段SAY 192 3 0 0 仅在cpDNA得到 .Southern杂交结果证实了片段SAY 192 3 0 0 的叶绿体来源 ,同时也表明在cms胞质中该片段所在区未发生插入、缺失等大的结构变异 .DNA序列测定结果表明 ,该片段为叶绿体ps1A1和ps1A2基因的部分序列 .图 6表 2参

The total DNAs from 15 allocytoplasmic or allonuclear sorghum species were analyzed by RAPD analysis, and a fragment SAY-19 2300 was amplified specifically from total DNAs of seven sorghum cultivars including 4 maintainers and 3 restorers with fertile (N) cytoplasm. PCR assays showed that this fragment was amplified from chloroplast (cp) DNA. The total DNA, mitochondrial (mt) DNA, and cpDNA were digested with EcoRI+HindIII, and hybridized with fragment SAY-19 2300 as probe. The hybridization signals appeared both in total DNA and cpDNA but not in mtDNA. No obviously and regularly polymorphic hybridization signals between the N- cytoplasm and sterile (S) cytoplasm were observed. Sequence analysis revealed this newly cloned fragment contained a portion of ps1A1 (1-2139bp) and ps1A2 (2165-2447bp) genes. Fig 6, Tab 2, Ref 19