Distribution of transfer RNA genes in thePisum sativum chloroplast DNA

Chu, NM; Shapiro,; Oishi, KK; Tewari, Κ. Κ.

doi:10.1007/bf02418752

Cited by 11 publications

(10 citation statements)

References 24 publications

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“…The structural organization of chloroplast DNA (ctDNA) has been well characterized [2,3,10] and genes for proteins, rRNAs and about 30 tRNAs have been identified and mapped in ctDNA [3,4,18,21,22]. The transcription of ctDNA has been studied using crude chloroplast extracts and recombinant DNA molecules containing rRNA, tRNA and protein genes [23].…”

Section: Introductionmentioning

confidence: 99%

Pea chloroplast topoisomerase I: purification, characterization, and role in replication

Nielsen

Tewari

1988

Plant Mol Biol

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A DNA-relaxing enzyme was purified 5 000-fold to homogeneity from isolated chloroplasts of Pisum sativum. The enzyme consists of a single polypeptide of 112 kDa. The enzyme was able to relax negatively supercoiled DNA in the absence of ATP. It is resistant to nalidixic acid and novobiocin, and causes a unit change in the linkage number of supercoiled DNA. The enzyme shows optimum activity at 37°C with 50 mM KCl and 10 mM MgCl2. From these properties, the enzyme can be classified as a prokaryotic type I topoisomerase.Using a partiall purified pea chloroplast DNA polymerase fraction devoid of topoisomerase I activity for in vitro replication on clones containing the pea chloroplast DNA origins of replication, a 2-6-fold stimulation of replication activity was obtained when the purified topoisomerase I was added to the reaction at 70-100 mM KCl. However, when the same reaction was carried out at 125 mM KCl, which does not affect DNA polymerase activity on calf thymus DNA but is completely inhibitory for topoisomerase I activity, a 4-fold drop in activity resulted. Novobiocin, an inhibitor of topoisomerase II, was not found to inhibit the in vitro replication of chloroplast DNA.

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Section: Introductionmentioning

confidence: 99%

Pea chloroplast topoisomerase I: purification, characterization, and role in replication

Nielsen

Tewari

1988

Plant Mol Biol

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“…We have reported the localization of tRNA genes in the restriction endonuclease map of pea ctDNA by hybridizing the 3 'end-labelled tRNAs with the appropriate restriction digests (6). These studies have identified eight different regions of ctDNA that contain tRNA genes.…”

Section: Resultsmentioning

confidence: 99%

“…The distribution of tRNA genes in pea ctDNA was analyzed by hybridizing 3' end-labelled tRNAs to the restriction digest of cloned recombinants (6). The nucleotide sequence analysis of those regions that hybridized with tRNAs has confirmed the location of tRNA genes and identified the specific tRNAs.…”

Section: Discussionmentioning

confidence: 99%

“…Plasmid DNA was obtained from the bacterial lysates by CsC1/EtBr density gradient centrifugation (34). Isolation of tRNAs from chloroplasts was carried out essentially by the method of Chu et al (6).…”

Section: Isolation Of Nucleic Acidsmentioning

confidence: 99%

“…Using this technique, 35 tRNA spots were separated and 21 tRNA genes mapped in spinach (11), 33 spots separated and 37 tRNA genes mapped in corn (44), 38 spots separated and 31 tRNA genes mapped in bean (31), 51 spots separated and 34 genes mapped in tobacco (1), and recently 33 spots separated and 31 tRNA genes mapped in wheat (30). Our recent studies on pea ct tRNA separated 36 tRNA spots (6). These tRNAs were eluted from the acrylamide gels and each hybridized to a series of recombinant pea ctDNA clones that had been found to contain tRNA genes by Southern hybridization.…”

Section: Introductionmentioning

confidence: 99%

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Nucleotide sequences of transfer RNA genes in the Pisum sativum chloroplast DNA

Shapiro

Tewari

1986

Plant Mol Biol

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Eight transfer RNA (tRNA) genes which were previously mapped to five regions of the Pisum sativum (pea) chloroplast DNA (ctDNA) have been sequenced. They have been identified as tRNA(Val)(GAC), tRNA(Asn)(GUU), tRNA(Arg)(ACG), tRNA(Leu)(CAA), tRNA(Tyr)(GUA), tRNA(Glu)(UUC), tRNA(His)(GUG), and tRNA(Arg)(UCU) by their anticodons and by their similarity to other previously identified tRNA genes from the chloroplast DNAs of higher plants or from E. gracilis. In addition,two other tRNA genes, tRNA(Gly) (UCC) and tRNA(Ile)(GAU), have been partially sequenced. The tRNA genes are compared to other known chloroplast tRNA genes from higher plants and are found to be 90-100% homologous. In addition there are similarities in the overall arrangement of the individual genes between different plants. The 5' flanking regions and the internal sequences of tRNA genes have been studied for conserved regions and consensus sequences. Two unusual features have been found: there is an apparent intron in the D-loop of the tRNA(Gly)(UCC), and the tRNA(Glu)(UUC) contains GATTC in its T-loop.

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Replication of Chloroplast DNA: Replication Origins, Topoisomerase I, and In Vitro Replication

1989

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SummaryReplication origins in pea chloroplast DNA (ctDNA) Using a partially purified pea chloroplast replication system, the supercoiled pCP12-7 and pCB1-12 recombinants served as highly active templates for DNA synthesis. Some smaller subclones of the A + T -rich D-Ioop region also show activity, including a 1.2 kbp BamHI-HindIII clone. A smaller 700 bp BamHI-BgnI clone shows very low activity suggesting that sequences outside the BgnI region are important for replication. Analysis of the DNA synthesised in vitro from these templates showed that full-length DNA was made, while other recombinants from other regions of the pea ctDNA showed no significant activity in vitro.Pea chloroplast topoisomerase I was purified 5000-fold to homogeneity, and was found to consist of a single polypeptide of 112,000 daltons. The enzyme catalyses an A TP-independent relaxation of negatively supercoiled DNA. It is resistant to nalidixic acid and novobiocin, and causes a unit change in the linkage number of supercoiled DNA. These properties support the identification of the pea chloroplast enzyme as a prokaryotic-like topoisomerase I.The role of topoisomerase I in the replication of ctDNA has been established by using a partially purified pea chloroplast replication system which lacks topoisomerase I activity, and recombinant templates which contain the pea ctDNA replication origins. A 2-6-fold stimulation of DNA synthesis resulted when the purified topoisomerase I was added to reactions at 80 mM KCl. When the reactions were carried out at 125 mM KC1, which is completely inhibitory for topoisomerase I but does not affect non-specific DNA polymerase activity, topoisomerase I addition caused no stimulation of activity. Novobiocin, an inhibitor of topoisomerase II, was found to have no effect on the in vitro replication of the ctDNA recombinants.Abbreviation: TPP, thymidine-5' -triphosphate. G. S. Singhal et al. (eds.), Photosynthesis

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Distribution of transfer RNA genes in thePisum sativum chloroplast DNA

Cited by 11 publications

References 24 publications

Pea chloroplast topoisomerase I: purification, characterization, and role in replication

Pea chloroplast topoisomerase I: purification, characterization, and role in replication

Nucleotide sequences of transfer RNA genes in the Pisum sativum chloroplast DNA

Replication of Chloroplast DNA: Replication Origins, Topoisomerase I, and In Vitro Replication

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