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1.
We applied human forensic techniques to the extraction of whole genomic DNA from processed wood samples to explore the possibility
of identifying an endangered tropical timber species by using DNA sequencing technology. High-yield and high-quality DNA samples
were obtained from 2 commercial wood and 3 herbarium samples. Large PCR fragments ranging from 500–800 bp were successfully
amplified from 2 chloroplast and 1 mitochondrial regions in all 5 samples, indicating limited degradation of the cytoplasmic
genomes. DNA extraction from stem wood taken from herbarium specimens appeared superior to that from stem wood with bark intact
or from leaf samples. DNA sequences from thetrn regions allowed for easy identification of the focal species based on GenBank Blast search. Little sequence variation was
observed in the 3 regions, with the mitochondrialcox3 region completely conserved. Extraction of high-quality and large intact DNA fragments makes dry wood materials amenable
to various DNA marker-based applications, including fingerprinting and historical approaches. By sampling stemwood, the wealth
of historical information housed in international herbaria can be explored with minimal damage to taxonomically important
features. 相似文献
2.
叶绿体基因组研究进展 总被引:14,自引:0,他引:14
作为植物细胞器的重要组成部分和光合作用的器官,叶绿体在生物进化的漫长历史中发挥了重要作用.伴随着生物技术的深入发展,人们发现叶绿体基因组结构和序列的信息在揭示物种起源、进化演变及其不同物种之间的亲缘关系等方面具有重要价值.与此同时,比核转化具有明显优势的叶绿体转化技术在遗传改良、生物制剂的生产等方面显示出巨大潜力,而叶绿体基因组结构和序列分析则是叶绿体转化的基石.基于叶绿体的这些重要作用,收集整理了有关的资料,从几个方面归纳了本领域最近的研究进展,希望能使读者对迅速发展的叶绿体基因组研究有更全面的了解,以及对叶绿体基因组在物种的进化、遗传、系统发育关系等方面的作用有更深刻的认识,同时也希望对叶绿体转化技术的研究和广泛应用产生积极作用. 相似文献
3.
Shengxin Chang Yankun Wang Jiangjie Lu Junyi Gai Jijie Li Pu Chu Rongzhan Guan Tuanjie Zhao 《PloS one》2013,8(2)
Determining mitochondrial genomes is important for elucidating vital activities of seed plants. Mitochondrial genomes are specific to each plant species because of their variable size, complex structures and patterns of gene losses and gains during evolution. This complexity has made research on the soybean mitochondrial genome difficult compared with its nuclear and chloroplast genomes. The present study helps to solve a 30-year mystery regarding the most complex mitochondrial genome structure, showing that pairwise rearrangements among the many large repeats may produce an enriched molecular pool of 760 circles in seed plants. The soybean mitochondrial genome harbors 58 genes of known function in addition to 52 predicted open reading frames of unknown function. The genome contains sequences of multiple identifiable origins, including 6.8 kb and 7.1 kb DNA fragments that have been transferred from the nuclear and chloroplast genomes, respectively, and some horizontal DNA transfers. The soybean mitochondrial genome has lost 16 genes, including nine protein-coding genes and seven tRNA genes; however, it has acquired five chloroplast-derived genes during evolution. Four tRNA genes, common among the three genomes, are derived from the chloroplast. Sizeable DNA transfers to the nucleus, with pericentromeric regions as hotspots, are observed, including DNA transfers of 125.0 kb and 151.6 kb identified unambiguously from the soybean mitochondrial and chloroplast genomes, respectively. The soybean nuclear genome has acquired five genes from its mitochondrial genome. These results provide biological insights into the mitochondrial genome of seed plants, and are especially helpful for deciphering vital activities in soybean. 相似文献
4.
Pervasive migration of organellar DNA to the nucleus in plants 总被引:1,自引:0,他引:1
A surprisingly large number of plant nuclear DNA sequences inferred to be remnants of chloroplast and mitochondrial DNA migration events were detected through computer-assisted database searches. Nineteen independent organellar DNA insertions, with a median size of 117 by (range of 38 to >785 bp), occur in the proximity of 15 nuclear genes. One fragment appears to have been passed through a RNA intermediate, based on the presence of an edited version of the mitochondrial gene in the nucleus. Tandemly arranged fragments from disparate regions of organellar genomes and from different organellar genomes indicate that the fragments joined together from an intracellular pool of RNA and/or DNA before they integrated into the nuclear genome. Comparisons of integrated sequences to genes lacking the insertions, as well as the occurrence of coligated fragments, support a model of random integration by end joining. All transferred sequences were found in noncoding regions, but the positioning of organellar-derived DNA in introns, as well as regions 5 and 3 to nuclear genes, suggests that the random integration of organellar DNA has the potential to influence gene expression patterns. A semiquantitative estimate was performed on the amount of organellar DNA being transferred and assimilated into the nucleus. Based on this database survey, we estimate that 3–7% of the plant nuclear genomic sequence files contain organellar-derived DNA. The timing and the magnitude of genetic flux to the nuclear genome suggest that random integration is a substantial and ongoing process for creating sequence variation.Correspondence to: J.L. Blanchard 相似文献
5.
In this work, we describe a method that allows the extraction of plant genomic DNA from frozen enzyme homogenates. To test if reliable DNA was recovered from allozyme extracts, we compared these results with that obtained from fresh frozen and dried material. Sequencing and microsatellite fragments were analyzed for samples from 5 plant families in order to check for chloroplast and nuclear DNA quality. High-molecular weight DNA was recovered from frozen homogenates, and it was found that the quality was independent of storage time, which varied from 1 to 9 years. High-quality PCR products along with sequences and microsatellite fragments were obtained for nearly all analyzed samples. Frozen protein homogenates from previous population studies could thus be used for further phylogeographical and phylogenetic analyses using DNA molecular tools. This simple method provides an opportunity to take advantage of stored cryopreserved samples. 相似文献
6.
Plants possess three major genomes, carried in the chloroplast, mitochondrion, and nucleus. The chloroplast genomes of higher plants tend to be of similar sizes and structure. In contrast both the nuclear and mitochondrial genomes show great size differences, even among closely related species. The largest plant mitochondrial genomes exist in the genus Cucumis at 1500 to 2300 kilobases, over 100 times the sizes of the yeast or human mitochondrial genomes. Biochemical and molecular analyses have established that the huge Cucumis mitochondrial genomes are due to extensive duplication of short repetitive DNA motifs. The organellar genomes of almost all organisms are maternally transmitted and few methods exist to manipulate these important genomes. Although chloroplast transformation has been achieved, no routine method exists to transform the mitochondrial genome of higher plants. A mitochondrial-transformation system for a higher plant would allow geneticists to use reverse genetics to study mitochondrial gene expression and to establish the efficacy of engineered mitochondrial genes for the genetic improvement of the mitochondrial genome. Cucumber possesses three unique attributes that make it a potential model system for mitochondrial transformation of a higher plant. Firstly, its mitochondria show paternal transmission. Secondly, microspores possess relatively few, huge mitochondria. Finally, there exists in cucumber unique mitochondrial mutations conditioning strongly mosaic (msc) phenotypes. The msc phenotypes appear after regeneration of plants from cell culture and sort with specific rearranged and deleted regions in the mitochondrial genome. These mitochondrial deletions may be a useful genetic tool to develop selectable markers for mitochondrial transformation of higher plants. 相似文献
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8.
A DNA extraction protocol for submerged pine logs was developed with the following properties: (i) high molecular weight DNA, (ii) PCR amplification of chloroplast and nuclear sequences, and (iii) high sequence homology to voucher pine specimens. The DNA extraction protocol was modified from a cetyltrimehtylammonium bromide (CTAB) protocol by adding stringent electrophoretic purification, proteinase K, RNAse, polyvinyl pyrrolidone (PVP), and Gene Releaser. Chloroplast rbcL (ribulose-1,5-bisphosphate carboxylase) could be amplified. Nuclear ribosomal sequences had >95% homology to Pinus taeda and Pinus palustris. Microsatellite polymorphism for PtTX2082 matched 2 of 14 known P. taeda alleles. Our results show DNA analysis for submerged conifer wood is feasible. 相似文献
9.
Yanti Rachmayanti Ludger Leinemann Oliver Gailing Reiner Finkeldey 《Plant Molecular Biology Reporter》2006,24(1):45-55
A successful DNA extraction from wood yielding appropriate DNA quality for PCR amplification allows molecular genetic investigations
of wood tissue. Genotypes, the origin of sampled material, and species can be identified based on an investigation of wood
if suitable information on genetic variation patterns within and among species is available. Potential applications are in
forensics and in the control of the timber and wood trade. We extracted DNA from wood of Dipterocarpaceae, a family that dominates
rainforests and comprises many important timber species in Southeast Asia. Several different DNA isolation techniques were
compared and optimized for wood samples from natural populations and from wood processing enterprises. The quality of the
DNA was tested by spectrophotometry, PCR amplification, and PCR inhibitor tests. An average DNA yield of 2.2 μg was obtained
per 50–100 mg of dried wood sample. Chloroplast DNA (cpDNA) regions of different length were amenable to PCR amplification
from the extracted DNA. Modification of DNA isolation techniques by the addition of polyvinylpyrrolidone (PVP) addition up
to 3.1% into lysis buffer reduced PCR inhibition effectively. In order to evaluate the extraction method, we analyzed leaves
and wood from the same tree by PCR amplification, genotyping and sequencing of chloroplast microsatellites. 相似文献
10.
N. W. Pirie 《BioEssays : news and reviews in molecular, cellular and developmental biology》1985,2(4):180-181
The existence and properties of the chloroplast genome were established by a combination of genetic methods which identified chloroplast mutations and placed them into a linear sequence or map; and by chemical methods, CsCl density gradient ultracentrifugation and base analysis, which identified non-nuclear DNA extracted from isolated chloroplasts. These studies, carried out in the 1950s and 1960s, primarily with Chlamydomonas, as well as parallel studies of mitochondrial DNA with yeast and Neurospora, laid the framework for distinguishing organelle and nuclear genomes. On this basis, the coding and regulatory functions of three genomes – nuclear, chloroplast, and mitochondrial – are being addressed in modern plant molecular biology. 相似文献
11.
D. Shimron-Abarbanell A. Breiman 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1991,83(1):71-80
Summary Scuttelar calli of Hordeum marinum readily and efficiently regenerate functional plants. In order to assess genetic variability among the regenerants we employed multiple analytic tools, which included molecular and biochemical assays. Total DNA extract from regenerated plants was digested with at least two restriction enzymes and hybridized to four nuclear and six mitochondrial coding sequences, in addition to one nuclear and three mitochondrial noncoding probes. SDS-PAGE analyses of hordein extracted from seeds of regenerated plants and activity assays of -amylase were also performed. The nuclear and mitochondrial genomes of 50 regenerated plants demonstrated relative stability when assessed with coding sequences and by biochemical analyses. However, the mitochondrial noncoding probes revealed one qualitative somaclonal variant characterized by a loss of a hybridizing fragment. Moreover, changes in the methylation patterns of the rRNA genes and the nontranscribed spacer were revealed in another regenerated plant. The albino plant regenerated was characterized by a loss of three chloroplast DNA BamHI fragments. 相似文献
12.
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14.
Background
The presence of chloroplast-related DNA sequences in the nuclear genome is generally regarded as a relic of the process by which genes have been transferred from the chloroplast to the nucleus. The remaining chloroplast encoded genes are not identical across the plant kingdom indicating an ongoing transfer of genes from the organelle to the nucleus.Scope
This review focuses on the active processes by which the nuclear genome might be acquiring or removing DNA sequences from the chloroplast genome. Present knowledge of the contribution to the nuclear genome of DNA originating from the chloroplast will be reviewed. In particular, the possible effects of stressful environments on the transfer of genetic material between the chloroplast and nucleus will be considered. The significance of this research and suggestions for the future research directions to identify drivers, such as stress, of the nuclear incorporation of plastid sequences are discussed.Conclusions
The transfer to the nuclear genome of most of the protein-encoding functions for chloroplast-located proteins facilitates the control of gene expression. The continual transfer of fragments, including complete functional genes, from the chloroplast to the nucleus has been observed. However, the mechanisms by which the loss of functions and physical DNA elimination from the chloroplast genome following the transfer of those functions to the nucleus remains obscure. The frequency of polymorphism across chloroplast-related DNA fragments within a species will indicate the rate at which these DNA fragments are incorporated and removed from the chromosomes.Key words: Stress, DNA transfer, organelles and nucleus, genome integration 相似文献15.
16.
New methods for better identification of timber geographical origin would constitute an important technical element in the forest industry, for phytosanitary certification procedures or in the chain of custody developed for the certification of timber from sustainably managed forests. In the case of the European white oaks, a detailed reference map of chloroplast (cp) DNA variation across the range exists, and we propose here to use the strong geographical structure, characterized by a differentiation of western vs. eastern populations, for the purpose of oak wood traceability. We first developed cpDNA markers permitting the characterization of haplotype on degraded DNA obtained from wood samples. The techniques were subsequently validated by confirming the full correspondence between genotypes obtained from living tissues (buds) and from wood collected from the same individual oak. Finally, a statistical procedure was used to test if the haplotype composition of a lot of wood samples is consistent with its presumed geographical origin. Clearly, the technique cannot permit the unambiguous identification of wood products of unknown origin but can be used to check the conformity of genetic composition of wood samples with the region of alleged origin. This could lead to major applications not only in the forest industry but also in archaeology or in palaeobotany. 相似文献
17.
Many applications in molecular ecology require the ability to match specific DNA sequences from single- or mixed-species samples with a diagnostic reference library. Widely used methods for DNA barcoding and metabarcoding employ PCR and amplicon sequencing to identify taxa based on target sequences, but the target-specific enrichment capabilities of CRISPR-Cas systems may offer advantages in some applications. We identified 54,837 CRISPR-Cas guide RNAs that may be useful for enriching chloroplast DNA across phylogenetically diverse plant species. We tested a subset of 17 guide RNAs in vitro to enrich plant DNA strands ranging in size from diagnostic DNA barcodes of 1,428 bp to entire chloroplast genomes of 121,284 bp. We used an Oxford Nanopore sequencer to evaluate sequencing success based on both single- and mixed-species samples, which yielded mean chloroplast sequence lengths of 2,530–11,367 bp, depending on the experiment. In comparison to mixed-species experiments, single-species experiments yielded more on-target sequence reads and greater mean pairwise identity between contigs and the plant species' reference genomes. But nevertheless, these mixed-species experiments yielded sufficient data to provide ≥48-fold increase in sequence length and better estimates of relative abundance for a commercially prepared mixture of plant species compared to DNA metabarcoding based on the chloroplast trnL-P6 marker. Prior work developed CRISPR-based enrichment protocols for long-read sequencing and our experiments pioneered its use for plant DNA barcoding and chloroplast assemblies that may have advantages over workflows that require PCR and short-read sequencing. Future work would benefit from continuing to develop in vitro and in silico methods for CRISPR-based analyses of mixed-species samples, especially when the appropriate reference genomes for contig assembly cannot be known a priori. 相似文献
18.
Successful amplification of rice chloroplast microsatellites from century-old grass samples from the park grass experiment 总被引:3,自引:0,他引:3
We report the successful amplification of microsatellite markers for the chloroplast genome from century-old samples of 2
grasses growing in the Park Grass Experiment (PGE):Anthoxanthum ordoratum andFestuca rubra. This opens the possibility of establishing a long-term genetic time series for the PGE, which began in 1856 and is believed
to be the oldest ecological experiment in existence. Although the plant samples used were not originally prepared or stored
with molecular analysis in mind, the hexadecyltrimethylammonium bromide (CTAB) method of DNA extraction was successfully used.
Obtained DNA was degraded but could be amplified by means of PCR. It produced bands around the expected size for chloroplast
microsatellite primers derived from rice. When sequenced, bands showed good homology with sequences from rice chloroplast
genomes listed in GenBank (accession No X15901). 相似文献
19.
Gilbertson L 《Trends in biotechnology》2003,21(12):550-555
Targeted insertion and the precise deletion of DNA from transgenic plant chromosomes increase the potential of plant biotechnology for commercial applications and basic research. The Cre–lox recombination system is one of the best characterized and most widely used systems for these purposes. Cre–lox has many applications, but it is primarily used for the controlled excision of DNA fragments, in particular selectable marker genes, from the nuclear and chloroplast genomes, and for the targeted insertion of DNA into specific sites in the nuclear genome. Recent developments, including regulated expression of cre and the creative use of wild-type and modified lox sites, have improved the potential of these applications. After almost 15 years of research and development in plants, the Cre–lox system continues to provide an efficient and precise tool for plant biotechnologists. 相似文献
20.
A polymorphic noncoding region of chloroplast DNA (cpDNA) was successfully amplified by the polymerase chain reaction (PCR) from various oak wood samples, including recent and more ancient (about 600-years-old) samples from different oak species. Adaptation of DNA isolation and amplification protocols was necessary to obtain this result. Polymorphisms useful to distinguish species or geographical origin of these samples could be scored through sequencing. These polymorphisms include one substitution and two microsatellite-type polymorphisms, due to a variable number of A/T repeats. Identical results were obtained independently in two separate laboratories. 相似文献