Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “DNA, Plant”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources
PubMed1990

Integration of Agrobacterium T-DNA into a tobacco chromosome: possible involvement of DNA homology between T-DNA and plant DNA.

We established tobacco tumour cell lines from crown galls induced by Agrobacterium. Restriction fragments containing T-DNA/plant DNA junctions were cloned from one of the cell lines, which has a single copy of the T-DNA in a unique region of its genome. We also isolated a DNA fragment that contained the integration target site from nontransformed tobacco cells. Nucleotide sequence analyses showed that the right and left breakpoints of the T-DNA mapped ca. 7.3 kb internal to the right 25 bp border and ca. 350 bp internal to the left border respectively. When the nucleotide sequences around these breakpoints were compared with the sequence of the target, significant homology was seen between the region adjacent to the integration target site and both external regions of the T-DNA breakpoints. In addition, a short stretch of plant DNA in the vicinity of the integration site was deleted. This deletion seems to have been promoted by homologous recombination between short repeated sequences that were present on both sides of the deleted stretch. Minor rearrangements, which included base substitutions, insertions and deletions, also took place around the integration site in the plant DNA. These results, together with previously reported results showing that in some cases sequences homologous to those in T-DNA are present in plant DNA regions adjacent to left recombinational junctions, indicate that sequence homology between the incoming T-DNA and the plant chromosomal DNA has an important function in T-DNA integration.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗
PubMed1975

On the question of the integration of exogenous bacterial DNA into plant DNA.

Extensive studies with pea, tomato, and barley failed to confirm the evidence presented by previous investigators for integration or replication of exogenously applied bacterial DNA in these plants. Labeled DNA of buoyant density in CsCl intermediate between that of high density donor bacterial DNA and of plant DNA was never observed with axenic plants. Intermediate peaks, similar to those used as evidence for recombination by earlier investigators, were observed only when the plants were contaminated with bacteria. Plant DNA prepared by a published procedure [Ledoux, L. & Huart, R. (1969) J. Mol. Biol. 43, 243-262] was found to be contaminated with unidentified impurities. Such DNA was partially protected from the action of DNase and produced aberrant banding patterns in CsCl after shearing. Much of the published evidence for integration of foreign DNA in plants is based upon experiments with plant DNA prepared by this procedure. We conclude that contamination is the likely explanation for what has been interpreted as evidence for integration.

Centrifugation, Density Gradient↗
PubMed1987

Integration of Agrobacterium tumefaciens transfer DNA (T-DNA) involves rearrangements of target plant DNA sequences.

The transfer DNA (T-DNA) mobilized into plant cells by Agrobacterium tumefaciens seems to integrate rather randomly into the plant genome. We analyzed a target site in the genome of Nicotiana tabacum before and after integration of a T-DNA. Clones presenting right and left T-DNA/plant DNA junctions were used as probes to identify and isolate a unique 1.8-kilobase EcoRI fragment corresponding to the plant DNA target site for a T-DNA insertion event. Comparison of the nucleotide sequences of the plant DNA portions of the T-DNA junction clones with the original plant DNA target revealed that several types of rearrangements resulted from insertion of the T-DNA. The most dramatic alteration was a 158-base-pair direct repeat of target plant sequences at the left and right T-DNA junctions. In addition, there were deletion and insertion events at the ends of the right and left copies of the 158-base-pair repeat. The variety of target-site rearrangements suggests that T-DNA insertion is a multistep process of recombination accompanied by local replicative and repair activities mediated by host-cell enzymes.

Journal Article↗
PubMed1978

DNA modifying enzymes of Agrobacterium tumefaciens: effect of DNA topoisomerase, restriction endonuclease, and unique DNA endonuclease on plasmid and plant DNA.

Extracts from Agrobacterium tumefaciens strain ID135 contain three enzymes that have been characterized and partially purified. The first enzyme, a DNA topoisomerase, appeared to relax only negatively twisted DNA. The second enzyme, Atu I, a type II restriction endonuclease, generated the identical DNA digestion pattern as EcoRII when several DNAs were used. The third enzyme, endonuclease A, showed a preference for superhelical DNAs as substrates. When plasmid pCK135DNA, obtained from the virulent strain IDI135 of A. tumefaciens, or plant DNA was exposed to the three enzymes, changes in DNA patterns were observed due to either conformational changes or digestion of the DNAs. These enzymes may function in vivo in the processing and incorporation of bacterial DNA in plant cells.

Animals↗
PubMed2002

Activation of antigen-presenting cells by immunostimulatory plant DNA: a natural resource for potential adjuvant.

Genomic DNA sequences (bacteria, insect, nematodes and molluscs) or synthetic oligodeoxynucleotides (ODN) containing unmethylated CpG motifs (CpG-DNA/ODN) are regarded as promising candidates for new medical adjuvants for their ability to stimulate the mammalian immune system and enhance immune responses to specific antigens. Here, we first report the immunostimulatory activity of total genomic DNA from two plants, Brassica chinensis L. and Zea may, the CpG methylation status of which is incomplete compared with E. coli DNA. These plant DNA can activate B cells to proliferate. Plant DNA promotes secretion of IL-12, and increases expression of MHC and costimulatory molecules by bone marrow-derived dendritic cells (BMDC). Plant DNA can also enhance antigen presentation capacity of BMDC and macrophages. When administrated in vivo, plant DNA can inhibit tumor growth in situ or metastasis in tumor-bearing mice. The immunostimulatory activity of plant DNA could be abolished by methylation. Our data showed that plant DNA can activate antigen-presenting cells (APC) including DC, macrophages and B cells, indicating that plant DNA is a new kind of potential adjuvant. Therefore, we conclude that plant DNA is another natural source of CpG-DNA, and that green plants may provide abundant resources for this potential medical adjuvant.

Adjuvants, Immunologic↗
PubMed2000

Plant DNA methyltransferases.

DNA methylation is an important modification of DNA that plays a role in genome management and in regulating gene expression during development. Methylation is carried out by DNA methyltransferases which catalyse the transfer of a methyl group to bases within the DNA helix. Plants have at least three classes of cytosine methyltransferase which differ in protein structure and function. The METI family, homologues of the mouse Dnmtl methyltransferase, most likely function as maintenance methyltransferases, but may also play a role in de novo methylation. The chromomethylases, which are unique to plants, may preferentially methylate DNA in heterochromatin; the remaining class, with similarity to Dnmt3 methyltransferases of mammals, are putative de novo methyltransferases. The various classes of methyltransferase may show differential activity on cytosines in different sequence contexts. Chromomethylases may preferentially methylate cytosines in CpNpG sequences while the Arabidopsis METI methyltransferase shows a preference for cytosines in CpG sequences. Additional proteins, for example DDM1, a member of the SNF2/SWI2 family of chromatin remodelling proteins, are also required for methylation of plant DNA.

DNA Methylation↗
PubMed2004

Sensitive PCR analysis of animal tissue samples for fragments of endogenous and transgenic plant DNA.

An optimized DNA extraction protocol for animal tissues coupled with sensitive PCR methods was used to determine whether trace levels of feed-derived DNA fragments, plant and/or transgenic, are detectable in animal tissue samples including dairy milk and samples of muscle (meat) from chickens, swine, and beef steers. Assays were developed to detect DNA fragments of both the high copy number chloroplast-encoded maize rubisco gene (rbcL) and single copy nuclear-encoded transgenic elements (p35S and a MON 810-specific gene fragment). The specificities of the two rbcL PCR assays and two transgenic DNA PCR assays were established by testing against a range of conventional plant species and genetically modified maize crops. The sensitivities of the two rbcL PCR assays (resulting in 173 and 500 bp amplicons) were similar, detecting as little as 0.08 and 0.02 genomic equivalents, respectively. The sensitivities of the p35S and MON 810 PCR assays were approximately 5 and 10 genomic equivalents for 123 bp and 149 bp amplicons, respectively, which were considerably less than the sensitivity of the rbcL assays in terms of plant cell equivalents, but approximately similar when the higher numbers of copies of the chloroplast genome per cell are taken into account. The 173 bp rbcL assay detected the target plant chloroplast DNA fragment in 5%, 15%, and 53% of the muscle samples from beef steers, broiler chickens, and swine, respectively, and in 86% of the milk samples from dairy cows. Reanalysis of new aliquots of 31 of the pork samples that were positive in the 173 bp rbcL PCR showed that 58% of these samples were reproducibly positive in this same PCR assay. The 500 bp rbcL assay detected DNA fragments in 43% of the swine muscle samples and 79% of the milk samples. By comparison, no statistically significant detections of transgenic DNA fragments by the p35S PCR assay occurred with any of these animal tissue samples.

Animal Feed↗
PubMed1988

RglB facilitated cloning of highly methylated eukaryotic DNA: the human L1 transposon, plant DNA, and DNA methylated in vitro with human DNA methyltransferase.

In vitro methylation of Bluescribe plasmid DNA (pBS) with human placental DNA methyltransferase to 6% 5-methylcytosine (mC) reduced transformation efficiencies in rglB+ host strains C600 and DS410 by almost 2 orders of magnitude. By contrast, the rglB- derivative of DS410 showed no reduction in transformation efficiency with methylation while the rglB- derivative of C600 was partially tolerant to methylation. Further, we show that the 1.8 kilobase (kb) and 1.2 kb KpnI fragments derived from the human L1 repeat have respectively 18.3% and 2.3% mC in vivo. Using these hyper- and hypo-methylated genomic segments ligated into the pBS plasmid, transformants with the highly methylated 1.8 kb L1 insert were recovered at 17 to 40 fold higher frequency with the rglB- host strains than with the rglB+ hosts. In addition, recombinant phage (lambda 2001) containing inserts of plant genomic DNA with 26.7% mC (from Petunia hybrida) when plated on rglB- hosts gave titres up to 222 times higher than on the rglB+ strains.

Cell Line↗
PubMed1995

Identification and analysis of a retinoblastoma binding motif in the replication protein of a plant DNA virus: requirement for efficient viral DNA replication.

Geminiviruses are plant DNA viruses with small genomes whose replication, except for the viral replication protein (Rep), depends on host proteins and, in this respect, are analogous to animal DNA tumor viruses, e.g. SV40. The mechanism by which these animal viruses create a cellular environment permissive for viral DNA replication involves the binding of a virally encoded oncoprotein, through its LXCXE motif, to the retinoblastoma protein (Rb). We have identified such a LXCXE motif in the Rep protein of wheat dwarf geminivirus (WDV) and we show its functional importance during viral DNA replication. Using a yeast two-hybrid system we have demonstrated that WDV Rep forms stable complexes with p130Rbr2, a member of the Rb family of proteins, and single amino acid changes within the LXCXE motif abolish the ability of WDV Rep to bind to p130Rbr2. The LXCXE motif is conserved in other members of the same geminivirus subgroup. The presence of an intact Rb binding motif is required for efficient WDV DNA replication in cultured wheat cells, strongly suggesting that one of the functions of WDV Rep may be the linking between viral and cellular DNA replication cycles. Our results point to the existence of a Rb-like protein(s) in plant cells playing regulatory roles during the cell cycle.

Amino Acid Sequence↗
PubMed2005

HPLC analysis of plant DNA methylation: a study of critical methodological factors.

HPLC analysis of nucleosides is important for determining total DNA methylation in plants and can be used to help characterise epigenetic changes during stress, growth and development. This is of particular interest for in vitro plant cultures as they are highly susceptible to genetic change. HPLC methodologies have been optimised for mammalian and microbial DNA, but not for plants. This study examines critical methodological factors in the HPLC analysis of plant DNA methylation using in vitro cultures of Ribes ciliatum. HPLC revealed that complete removal of RNA from plant DNA extractions is difficult using RNase (A and T1) digestions and LiCl precipitation. This suggests that base analysis should be avoided when using these RNA removal techniques, as bases from residual RNA fragments will inflate peak areas for DNA-derived bases. Nucleoside or nucleotide analysis is therefore recommended as a more suitable option as RNA and DNA constituents can be readily separated. DNA digestion was also a critical factor as methylation was under-estimated following incomplete nuclease digestion and over-estimated following incomplete phosphatase digestion. The units of enzyme required for complete DNA digestion was optimised and found to be 20-200 times less for nuclease P1 and 15 times less for alkaline phosphatase as compared with previous protocols. Digestion performance was conveniently monitored using marker peaks that indicate incomplete digestion products. This study identifies critical components of HPLC analysis and offers a comprehensive guide for the stringent analysis of DNA methylation in plants.

Chromatography, High Pressure Liquid↗
PubMed1997

Molecular cloning of the cDNA for the catalytic subunit of plant DNA polymerase alpha and its cell-cycle dependent expression.

BACKGROUND: DNA polymerase alpha has been studied in considerable detail in yeast and animals. Genetic and biochemical analyses reveal that this enzyme is composed of a heterotetramer and is necessary for replicon initiation and primer synthesis in lagging strand synthesis. In spite of the fact that modes of DNA replication in plants seem to be similar to those in other eukaryotes, very little is known about the biochemical components that participate in DNA replication of plants, including DNA polymerases. RESULTS: Using a 561-base pair DNA fragment, obtained by polymerase chain reaction amplification from a rice cDNA library as a probe, we isolated and sequenced a cDNA homologous to the cDNA for the catalytic subunit of rice DNA polymerase alpha. The encoded polypeptide has extensive homology with the catalytic subunit of DNA polymerase alpha from several species. Furthermore, when the cDNA was expressed in eukaryotic transcription/translation systems, the protein products showed DNA polymerase activity which was inhibited by a monoclonal antibody specific for DNA polymerase alpha. Using RNA gel blot analysis, we found that the levels of mRNA of the catalytic subunit of this enzyme is regulated during the cell-cycle in plant cells. CONCLUSION: This is the first report which describes the cDNA cloning of plant DNA polymerase. We conclude that the principal features of the DNA polymerase alpha catalytic subunit are conserved in plants.

Amino Acid Sequence↗
PubMed2006

Release and leaching of plant DNA in unsaturated soil column.

Little information is available on the process of DNA release from plants and neither is there much information to be found regarding DNA transport in the vadose zone. Unsaturated soil columns were used to examine the release and transport of DNA content in the leaf of tomato variety Palmiro, which was introduced into the soil columns after being dried at 35 degrees C for 3 days. Soil columns were leached with sterile water at a rate of 0.5mLh(-1) for 104 days. DNA from column leachate water was extracted and analysed. Both quantitative and qualitative measurements of extracted DNA were taken. Tomato gene sequences were detected using PCR amplification based on tomato specific 18S primers. The concentration of total DNA in soil column leachate water ranged from 6.7 to 50.4microgL(-1). Genomic analysis using agarose gel electrophoresis showed degradation of DNA after its passage through unsaturated soil columns. The presence of tomato genes in the leachate water of soil columns suggests a potential release of tomato DNA from leaves and potential transport of DNA over considerable distances in water-unsaturated soil. Consequently, transport of plant DNA in vadose zone is indicated, and there is a risk that the DNA may reach the groundwater.

DNA, Plant↗
PubMed2002

Impact of feed processing and mixed ruminal culture on the fate of recombinant EPSP synthase and endogenous canola plant DNA.

The impact of feed processing and in vitro ruminal cultures on the persistence of recombinant and canola-specific endogenous DNA was studied using various canola substrates (whole seed, cracked seed, meal and diet). For both, parental and genetically modified substrates, ribulose-1,5-bisphosphate carboxylase/oxygenase gene was amplifiable up to varying time points. Persistence of recombinant DNA, encoding 5-enolpyruvylshikimate-3-phosphate synthase (1,363 bp) was detected up to 8 h for meal and 4 h for mixed diet. Upon processing of canola, DNA large enough to contain intact plant genes remains. In an in vitro environment, plant DNA was rapidly degraded upon its release into rumen fluid.

3-Phosphoshikimate 1-Carboxyvinyltransferase↗
PubMed1982

DNA from the A6S/2 crown gall tumor contains scrambled Ti-plasmid sequences near its junctions with plant DNA.

The A6S/2 tumor incited on tobacco by Agrobacterium tumefaciens harboring the octopine-type A6 Ti plasmid contains one insert of Ti-plasmid sequences (the T DNA). This 13 kb insert is derived from a colinear sequence in the Ti plasmid (the T region) and becomes attached to plant DNA in the nucleus of the host cell. We have determined the DNA sequence encompassing the left end of the T region of the A6 Ti plasmid and the corresponding portion of the A6S/2 T DNA. The two sequences are identical for at least 806 bp. To the left of the divergence point, the tumor contains five partially overlapping sequences that are direct or inverted repeats of sequences to the right of the divergence point. The Ti plasmid contains only the right member of each of these repeats. We have also performed heteroduplex studies that indicate that this T DNA has a 520 bp inverted repeat of an internal sequence at the right end near its junction with plant DNA. The repeated sequences near the ends of the T DNA resemble the repeats of adenovirus type 12 sequences found near its junction with host DNA. We discuss data suggesting that the 23 bp to the immediate right of the divergence point of the A6 left junction form a site important in some step in the transfer of T-region DNA from the bacteria to the plant.

Base Sequence↗
PubMed1988

Supercoils in plant DNA: nucleoid sedimentation studies.

Plant nuclei have been studied with respect to the three-dimensional structure of DNA. Nucleoids derived from nuclei by non-ionic detergent and high salt treatment were analysed by sedimentation in a series of sucrose gradients containing increasing amounts of the intercalating agent ethidium bromide. In addition the nucleoid sedimentation behaviour was investigated following gamma irradiation. The results show that plant DNA is supercoiled, as is the DNA from the other eukaryotes studied, and contains approximately the same concentration of superhelical turns but probably relatively fewer DNA superhelical loops. The plant nuclear populations in all cases studied give rise to two distinct nucleoid bands. These have been characterized by electron microscopy and by their DNA and protein content. The possible origin of the two bands is discussed.

Cell Nucleus↗
PubMed1981

Determination of 5-methylcytosine from plant DNA by high-performance liquid chromatography.

The relative amounts of the five nucleosides (deoxycytidine, 5-methyldeoxycytidine, deoxyadenosine, deoxyguanosine and thymidine) in the DNA of nine plant species, one plant satellite DNA, and one animal species were determined by high performance liquid chromatography. The method allows the clean separation of the nucleosides from 10 microgram samples with 15 min. The following values for the proportion of methylated cytosines among all cytosines were obtained: Lobularia maritima 18.5%, Nicotiana tabacum 32.6%, Pisum sativum 23.2%, Rhinanthus minor 29.2%, Sinapsis alba 12.2%, Vicia faba 30.5%, Viscum album 23.2%, Cymbidium pumilum 18.8%, Cymbidium pumilum AT-rich satellite DNA 15.8%, Triticum aestivum 22.4%. DNA of an animal, the gerbil, Meriones unguiculatus, had a methylation percentage of 3.1%. An estimate of the GC content based on the buoyant density of DNA tends to be lower than the actual value, an estimate based on the melting temperature tends to be higher. This supports the finding by other authors that DNA methylation decreases the buoyant density and may increase the melting temperature at high m5C concentration.

5-Methylcytosine↗
PubMed2006

Degradation of plasmid and plant DNA in water microcosms monitored by natural transformation and real-time polymerase chain reaction (PCR).

Extracellular DNA exists in the environment and can be taken up by competent bacterial cells, leading to horizontal gene transfer. The persistence of extracellular plasmid and plant DNA in water microcosms was monitored in this study. Water samples were two groundwater (GW1 and GW2) and one river water (RW) samples. Three treatments included: (1) intact, (2) 0.22 microm filter-sterilized, and (3) autoclaved water. DNA from a plasmid (pNS1) and a transgenic Bt (Bacillus thuringiensis) corn line, both carrying a neomycin phosphotransferase gene (nptII gene) conferring kanamycin and neomycin resistance, was inoculated into the microcosms at 0.4 and 0.8 microg/ml, respectively. By monitoring its ability to transform a competent Pseudomonas stutzeri strain harboring plasmid pMR7 (P. stutzeri pMR7), plasmid DNA was degraded to undetectable levels in the intact and/or filter-sterilized water treatments within 48-96 h in GW1, GW2, and RW. Meanwhile, plasmid DNA persisted in the autoclaved treatment throughout the entire incubation period. For plant DNA, a highly sensitive real-time PCR method using SYBR Green I was developed to monitor the degradation dynamics of the nptII gene carried by the transgenic corn line in the microcosms. The results showed that the concentration of plant DNA was reduced by two orders of magnitude (from 0.8-0.008 microg/ml) within 96 h in the intact and filter-sterilized treatments of GW1, GW2, and RW, in contrast to its persistence in the autoclaved treatment. In addition, no kanamycin resistant (Km R) transformants were detected from in situ transformation of P. stutzeri pMR7 with plasmid pNS1 DNA.

Bacillus thuringiensis↗