绿藻Heterochlorella luteoviridis SAG211-2a叶绿体16S rRNA基因研究

Study on 16S rRNA gene structure in the Chloroplast of Heterochlorella luteoviridis SAG211-2a

为了研究绿藻Heterochlorella luteoviridis SAG211-2a叶绿体16S rRNA基因中内含子和双发夹结构在rRNA成熟过程中是否被切除,以来自德国歌德堡大学藻种库的绿藻藻株SAG 211-2a为材料,提取其基因组DNA作为模板,PCR扩增并测序叶绿体16S rRNA基因,与同源物种的16S rRNA基因进行比对;并提取SAG211-2a的总RNA,用一管法RT-PCR法扩增16S rDNA基因成熟转录子后,克隆至pMD19-T表达载体上,转化大肠杆菌DH5α,提取质粒并测序。结果表明,绿藻SAG211-2a叶绿体16S rRNA基因中存在1个长度为46 bp的双发夹结构(DHE)和1个长度为537 bp的Ⅰ类内含子,内含子在rRNA成熟过程中被切除,而双发夹结构却保留在rRNA中,从而验证了双发夹结构在rRNA成熟过程中没有被切除。

In order to study whether the intron and double-hairpin structure in the Chloroplast 16S rRNA gene of Heterochlorella luteoviridis SAG211-2a were removed during the maturation of rRNA, H. luteoviridis SAG211-2a coming from the algae collection of Gothenburg University in Germany was used as material. The 16S rRNA gene was amplified by PCR using the extracted H. luteoviridis SAG211-2a DNA as template, and was compared with the 16S rRNA from homologous species. Then, the 16S rRNA gene mature transcript was amplified by one step of RT-PCR method using the total RNA extracted from H. luteoviridis as template and was inserted into expression vector pMD19-T, then was transformed to E. coli DH5α. Plasmids were extracted and sequenced. The results showed that the Chloroplast 16S rRNA gene of H. luteoviridis SAG211-2a had a double-hairpin structure （DHE） of 46 bp and a Group I intron of 537 bp. The Group I intron was removed in the rRNA maturation, and the double-hairpin structure was retained in the rRNA. Thus verified that the double-haipin structure was not removed in the rRNA maturation.