Briat J F, Dron M, Loiseaux S, Mache R
Nucleic Acids Res. 1982 Nov 11;10(21):6865-78. doi: 10.1093/nar/10.21.6865.
A cloned fragment of spinach chloroplast DNA carrying 140 bp of the 16S rRNA gene and 691 bp upstream this gene has been analysed by DNA sequencing, by in vitro transcription, by S1 mapping with chloroplast RNAs and purified 16S rRNA from 30S ribosomal subunits. A tRNAVal gene has been located between the position 394 and 465. Crude chloroplast RNA polymerase has been purified by heparin sepharose chromatography of a 80 000 g supernatant from pure lysed spinach plastids and used to transcribe the cloned Bg1 II-Pvu II DNA fragment. Four in vitro transcripts of about 830, 550, 350 and 260 bases were obtained whatever RNA polymerase used: the chloroplast or the E. coli enzyme. The transcripts of 550 and 260 bases are initiated by ATP. S1 mapping with in vivo chloroplasts RNAs on 5' labelled separated strands from Bg1 II-Pvu II fragments indicates 2 protected DNA fragments respectively of 140 and 260 bases on the strand which codes for rRNAs and possibly one protected DNA fragment of 550 bases on the other strand. The start site of the 260 bases transcript might correspond to the initiation site of transcription of the rRNA genes. The possibility that the 550 bases transcription of the non coding strand for rRNA genes corresponds to the beginning of a mRNA is discussed.
对菠菜叶绿体DNA的一个克隆片段进行了分析，该片段含有16S rRNA基因的140bp以及该基因上游的691bp，分析方法包括DNA测序、体外转录、用叶绿体RNA进行S1作图以及从30S核糖体亚基中纯化16S rRNA。已确定一个tRNAVal基因位于394至465位之间。通过对纯裂解菠菜质体80000g上清液进行肝素琼脂糖层析，纯化了粗制的叶绿体RNA聚合酶，并用于转录克隆的Bg1 II - Pvu II DNA片段。无论使用叶绿体RNA聚合酶还是大肠杆菌酶，都获得了大约830、550、350和260个碱基的四种体外转录本。550和260个碱基的转录本由ATP起始。用体内叶绿体RNA对Bg1 II - Pvu II片段5'标记的单链进行S1作图，结果表明，在编码rRNA的链上分别有140和260个碱基的2个受保护的DNA片段，在另一条链上可能有一个550个碱基的受保护的DNA片段。260个碱基转录本的起始位点可能对应于rRNA基因的转录起始位点。文中讨论了rRNA基因非编码链550个碱基的转录对应于一个mRNA起始的可能性。
