摘要
本文报道对一个衰老下调基因的鉴定 .从绿色和正在衰老的紫萍叶状体组织中分别提取总 RNA,然后使用一个 oligo-d T引物进行逆转录合成 c DNA.c DNA用 oligo-d T和简并引物进行 PCR扩增 ,然后进行琼脂糖凝胶电泳 .电泳后发现 ,一条 c DNA带存在于由绿色组织而不存在于由正在衰老的组织所得到的样品中 .将这一 c DNA回收 ,克隆进 T-vector并且进行了测序 .Northern blot分析表明 ,该 c DNA对应基因的表达受衰老调节而减少 .根据第一个 c DNA序列设计了一个特异性引物 ,进行 RT-PCR扩增 ,并且克隆到另外一个部分 c D-NA.测序结果表明 ,第二个克隆具有与第一个 c DNA相重叠的 40 bp的片段 .将第一个 c DNA序列与删去 40 bp重叠区的第二个 c DNA序列连接 ,我们得到一个 3 2 2个核苷酸的 c DNA3′端部分序列 .用 BLASTN程序将该序列翻译后与 NCBI non-redundant database中已知的蛋白进行比较 ,结果表明所克隆的 c DNA代表的基因与光呼吸途径的丝氨酸 -乙醛酸氨基转移酶编码基因有很高的相似性 .
The priliminary identification of a gene down regulated by senescence is reported in this paper. Total RNA was isolated from green and senescing duckweed frond tissues and reverse transcribed into cDNAs using an oligo dT primer. After amplification with PCR using the oligo dT primer and degenerate primers, the cDNAs were electrophoresed on an agarose gel. One cDNA band was found to be present in the sample prepared from green but not from senescing tissues. This cDNA was recovered, cloned into a T vector and sequenced. Northern blot analysis showed that the expression of the gene corresponding to the cDNA was down regulated by senescence. Another partial cDNA was also amplified by RT PCR using a primer designed from the first clone. Sequencing of the second clone shows that it contains an overlap part of 40bp with the first cDNA. By melding these two sequences we got a 3′ partial cDNA sequence of 322 nucleotides. This sequence was translated and compared with known proteins in the NCBI non redundant database using the BLASTN program. The result shows that the gene corresponding to the cDNAs cloned has a very high similarity to genes encoding serine glyoxylate aminotransferase, an enzyme involved in photorespiration.
出处
《南开大学学报（自然科学版）》
CAS
CSCD
北大核心
2004年第1期69-74,共6页
Acta Scientiarum Naturalium Universitatis Nankaiensis
基金
Suppoted by NSFC( 3 9970 0 77)
关键词
衰老
基因表达
丝氨酸-乙醛酸氨基转移酶
紫背浮萍
senescence
gene expression
serine glyoxylate aminotransferase
Spirodela polyrrhiza
