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1.
The chloroplast genome ofChlamydomonas reinhardtii has been transformed with a chimeric gene consisting of the chloroplastatpA promoter and the bacterial gene for aminoglycoside adenine transferase (aadA). TheatpA-aadA cassette has been placed within the chloroplast DNAEcoRI restriction enzyme fragment 14, or within the chloroplastBamH1 fragment 10. The chimeric constructs were introduced into the chloroplast by particle bombardment. Integration of the cassette into chloroplast DNA then occurred via homologous recombination of sequences flanking the cassette with their corresponding chloroplast sequences. We demonstrate that the chloroplastatpA promoter inatpA-aadA routinely recombines with its endogenous counterpart, resulting in heteroplasmic chloroplast DNA populations that may persist for many generations. The heterologous gene does not require a 3 inverted repeat sequence for its expression. TheatpA-aadA gene copy number, which is dictated here by its position in the chloroplast genome, is proportional to the steady state level ofatpA-aadA mRNA. However, neither genomic position, gene copy number, or mRNA level have a significant effect on cellular resistance to spectinomycin, nor activity of theaadA gene productin vitro. These results suggest that, in the case ofaadA, the limiting step for expression of this gene is at the translational or post-translational level. TheatpA-aadA cassette should prove a useful model for future studies on the maintenance and expression of heterologous genes inC. reinhardtii chloroplasts. 相似文献
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The location on the wheat chloroplast DNA map and the nucleotide sequences of the genes coding for tRNA
GCC
Gly
(trnG-GCC), tRNA
GUC
Asp
(trnD-GUC) and tRNA
GCA
Cys
(trnC-GCA) have been determined. These three genes are located in the large single copy region of the chloroplast genome, about
half-way between one of the inverted repeats and the gene for the α subunit of ATP synthase. They are located on two Bam H1
fragments, called B6 and B18 by Bowmanet al. (1), which are separated by about 450 bp and which were cloned in our laboratory to allow sequencing. ThetrnD-GUC andtrnC-GCA sequences show 98.6 and 89% homology, respectively, with the corresponding spinach chloroplast tRNA genes sequences
(2), which are the only other higher plant chloroplasttrnD-GUC andtrnC-GCA sequenced so far, while no othertrnG-GCC sequence has been published. ThetrnG-GCC sequence shows only 58% homology with the corresponding gene sequence inEuglena chloroplasts (3). 相似文献
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Genomic, proteomic, phylogenetic and evolutionary aspects of a novel gene encoding a putative chloroplast-targeted sulfate permease of prokaryotic origin in the green alga Chlamydomonas reinhardtii are described. This nuclear-encoded sulfate permease gene (SulP) contains four introns, whereas all other known chloroplast sulfate permease genes lack introns and are encoded by the chloroplast genome. The deduced amino acid sequence of the protein showed an extended N-terminus, which includes a putative chloroplast transit peptide. The mature protein contains seven transmembrane domains and two large hydrophilic loops. This novel prokaryotic-origin gene probably migrated from the chloroplast to the nuclear genome during evolution of C. reinhardtii. The SulP gene, or any of its homologues, has not been retained in vascular plants, e.g. Arabidopsis thaliana, although it is encountered in the chloroplast genome of a liverwort (Marchantia polymorpha). A comparative structural analysis and phylogenetic origin of chloroplast sulfate permeases in a variety of species is presented. 相似文献
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Summary The circular chloroplast DNA from three species of plants in the taxonomic family Leguminosae were examined using electron microscopic techniques and restriction endonuclease digestion. Chloroplast DNAs from chickpea (Cicer arietinum), mung bean (Vigna radiata), and soy bean (Glycine max) were found to range in size from 119–151 kilobase pairs by contour length measurements. Sizes of the chloroplast DNAs have been further confirmed using different restriction endonucleases. Two of the chloroplast DNAs examined, soy bean and mung bean, contain a region approximately 15.9–18% of their monomer length that is repeated in reverse polarity. This repeated region separates a small unique region that ranges in size from 18.75–20.4 kilobase pairs and a large unique region that ranges in size from 73.4–85 kbp. This feature was not found in the chloroplast DNA of chickpea. R-loop hybridizations performed using chloroplast ribosomal RNAs demonstrate that the two ribosomal gene sets of the mung been and soy bean are arranged in inverted orientation within this repeated region. In contrast, the chickpea chloroplast DNA posesses a single ribosomal RNA gene set in the circular molecule. In all three chloroplast DNAs examined, the genes encoding the chloroplast 23S and 16S ribosomal RNA genes are separated by a spacer region which ranges in size from 2.2 to 2.48 kbp. 相似文献
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Klaus Valentin 《Molecular & general genetics : MGG》1993,236(2-3):245-250
Summary Partial sequence analysis of the plastid DNA (ptDNA) from a red alga, Antithamnion sp., revealed the presence of a homologue to the Escherichia coli SecA gene as well as two open reading frames (ORF 510, ORF 179). In addition a sec Y homologue has been detected on the plastid genome by heterologous hybridization. None of these genes has been found in completely sequenced chlorophytic plastid genomes. SecA and secY gene copies were also detected in the ptDNA of a chromophytic alga, indicating that secAY may be ubiquitous in rhodophytes and chromophytes. The significance of these findings for the evolution of plastid genomes and the thylakoid protein import mechanism is discussed. 相似文献
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小麦叶绿体蛋白质编码基因RNA编辑位点的测定及与返白现象的关系 总被引:1,自引:0,他引:1
RNA编辑是一种转录后基因加工修饰现象,广泛存在于高等植物细胞器中。已有研究表明,RNA编辑与植物发生白化或者黄化有关。通过PCR、RT-PCR及测序的方法,对具有阶段性白化特性的小麦(Triticum aestivum)返白系FA85及其野生型矮变一号(Aibian 1)的叶绿体蛋白质编码基因RNA编辑位点进行了测定,在14个基因上发现了26个编辑位点。有5个编辑位点在2个株系之间存在编辑效率的差异,且这些差异的位点均位于编码叶绿体RNA聚合酶的基因上,其中3个位点编辑前后对应的蛋白质二级结构可能有差异。对2个株系叶绿体中PEP、NEP及PEP、NEP共同依赖基因转录水平的检测显示,除psbA和clpP外,其它基因在小麦返白系中的转录水平均有不同程度的下降。这种转录水平的显著下降及叶绿体RNA聚合酶基因上RNA编辑位点编辑效率的改变,可能与小麦返白系叶片的返白有关。 相似文献
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A phylogenetic reconstruction of eight species of the genusBrachypodium P. Beauv. (Poaceae) has been obtained combining sequence data from the chloroplastndhF gene and the nuclear ITS, and using five representatives of tribes Triticeae (Secale), Poeae (Lolium), Meliceae (Melica, Glyceria) and Oryzeae (Oryza) as out-groups. Similar numbers of informative substitutions for the ingroup species were provided by both the 3 region of the chloroplastndhF gene and the nuclear ITS region. The Mediterranean annualBrachypodium distachyon appears to be the basal lineage, followed by the divergence of the New World non-rhizomatousB. mexicanum, which antedates the separation of a core of six European and Eurosiberian rhizomatous perennials (Brachypodium arbuscula, B. retusum, B. rupestre, B. phoenicoides, B. pinnatum, andB. sylvaticum). The evolutionary reconstruction based on sequences of the chloroplast and the nuclear genomes is congruent with topologies obtained from analysis of RAPD data. 相似文献
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Hans J. Bohnert Karl H. J. Gordon Edwin J. Crouse 《Molecular & general genetics : MGG》1980,179(3):539-545
Summary Labelled chloroplast rRNAs from Spinacia oleracea were hybridized to restriction endonuclease digests of chloroplast DNA from Oenothera hookeri and Euglena gracilis, to mitochondrial DNA of Acanthamoeba castellanii, and to DNA of the E. coli rrn B operon in the transducing phage lambda rifd18. The degree of homology is greatest for the 16S rRNA gene. Greater than 90% occurs between the two higher plant genes, 80% homology to the lower plant gene, 60%–70% homology to the bacterial gene, and 20% homology to the mitochondrial gene. The degree of hybridization varied considerably for the 23S and the 5S rRNA genes. Very high homology exists between the two higher plant genes, only about 50% homology for both the Euglena and bacterial genes, and no significant homology for the mitochondrial genes. These results show that any chloroplast (or E. coli) rRNA may be used as a probe to identify rRNA genes in other ctDNAs.Two RNA populations, each enriched for a different ctDNA-encoded mRNA, proved useful in the location of these genes on both higher plant ctDNAs. No significant hybridization was obtained using these probes to the Euglena ctDNA which seems to be too distantly related.Abbreviations Md
megadalton, 106 dalton
- bp, kbp
base pair, kilo base pair
- SSC
Standard saline citrate, 1 times SSC is 0.15M sodium, chloride, 0.015 M trisodium citrate, pH, 6.8
- mtDNA
mitochondrial DNA
- ctDNA
chloroplast DNA
- ctrRNA
chloroplast ribosomal RNA 相似文献
15.
V. Goremykin K. I. Hirsch-Ernst S. Wölfl F. H. Hellwig 《Plant Systematics and Evolution》2003,242(1-4):119-135
The nucleotide sequence of the complete chloroplast genome of a basal angiosperm, Calycanthus
fertilis, has been determined. The circular 153337 bp long cpDNA is colinear with those of tobacco, Arabidopsis and spinach. A total of 133 predicted genes (115 individual gene species, 18 genes duplicated in the inverted repeats) including 88 potential protein-coding genes (81 gene species), 8 ribosomal RNA genes (4 gene species) and 37 tRNA genes (30 gene species) representing 20 amino acids were identified based on similarity to their homologs from other chloroplast genomes. This is the highest gene number ever registered in an angiosperm plastome. Calycanthus fertilis cpDNA also contains a homolog of the recently discovered mitochondrial ACRS gene. Since no gene transfer from mitochondria to the chloroplast has ever been documented, we investigated the evolutionary affinity of this gene in detail. Phylogenetic analysis of the protein-coding subset of the plastome suggests that the ancient line of Laurales emerged after the split of the angiosperms into monocots and dicots.
Calycanthus fertilis Walter var. ferax (Michy.) Rehder is a synonym of C. floridus L. var. glaucus (Willd.) Torr. & A. Gray.Data deposition: The sequence reported in this paper has been deposited in the EMBL database (accession no. AJ428413). 相似文献
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Scott M. Newman Nicholas W. Gillham Elizabeth H. Harris Anita M. Johnson John E. Boynton 《Molecular & general genetics : MGG》1991,230(1-2):65-74
Summary We have developed an efficient procedure for the disruption of Chlamydomonas chloroplast genes. Wild-type C. reinhardtii cells were bombarded with microprojectiles coated with a mixture of two plasmids, one encoding selectable, antibiotic-resistance mutations in the 16S ribosomal RNA gene and the other containing either the atpB or rbcL photosynthetic gene inactivated by an insertion of 0.48 kb of yeast DNA in the coding sequence. Antibiotic-resistant transformants were selected under conditions permissive for growth of nonphotosynthetic mutants. Approximately half of these transformants were initially heteroplasmic for copies of the disrupted atpB or rbcL genes integrated into the recipient chloroplast genome but still retained photosynthetic competence. A small fraction of the transformants (1.1% for atpB; 4.3% for rbcL) were nonphotosynthetic and homoplasmic for the disrupted gene at the time they were isolated. Single cell cloning of the initially heteroplasmic transformants also yielded nonphotosynthetic segregants that were homoplasmic for the disrupted gene. Polypeptide products of the disrupted atpB and rbcL genes could not be detected using immunoblotting techniques. We believe that any nonessential Chlamydomonas chloroplast gene, such as those involved in photosynthesis, should be amenable to gene disruption by cotransformation. The method should prove useful for the introduction of site-specific mutations into chloroplast genes and flanking regulatory sequences with a view to elucidating their function. 相似文献
19.
Dr. Wipa Chungjatupornchai 《Current microbiology》1990,21(5):283-288
CyanobacteriumSynechocystis PCC6803 was used as a model system for a prolonged delivery of insecticidal crystal proteins ofBacillus thuringiensis subsp.israelensis in the water surface. Thebt8 gene, encoding a 128 kDa (Bt8) mosquitocidal protein, and the28kd gene, encoding a 28 kDa cytotoxic protein, were integrated into the cyanobacterial chromosome. The genes were expressed under the control of thepsbA promoter, derived from the tobacco chloroplast genome. The Bt8 protein produced by the cyanobacterium was toxic to mosquito larvae. The28kd gene expression in the cyanobacterium was very low, partly owing to the low level of steady state mRNA. With the same system, it was demonstrated that the herbicide-resistance genebar could be used as a new selectable marker in cyanobacterial transformation experiments. 相似文献
20.
BEL1‐LIKE HOMEODOMAIN 11 regulates chloroplast development and chlorophyll synthesis in tomato fruit
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Lanhuan Meng Zhongqi Fan Qiang Zhang Cuicui Wang Ying Gao Yikang Deng Benzhong Zhu Hongliang Zhu Jianye Chen Wei Shan Xueren Yin Silin Zhong Donald Grierson Cai‐Zhong Jiang Yunbo Luo Da‐Qi Fu 《The Plant journal : for cell and molecular biology》2018,94(6):1126-1140