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1.
David N. Kuhn Antonio Figueira Uilson Lopes Juan Carlos Motamayor Alan W. Meerow Kathleen Cariaga Barbie Freeman Donald S. LivingstoneIII Raymond J. Schnell 《Tree Genetics & Genomes》2010,6(5):783-792
The seeds of Theobroma cacao (cacao) are the source of cocoa, the raw material for the multi-billion dollar chocolate industry. Cacao’s two most important
traits are its unique seed storage triglyceride (cocoa butter) and the flavor of its fermented beans (chocolate). The genome
of T. cacao is being sequenced, and to expand the utility of the genome sequence to the improvement of cacao, we are evaluating Theobroma grandiflorum, the closest economically important species of Theobroma for its potential use in a comparative genomic study. T. grandiflorum differs from cacao in important agronomic traits such as flavor of the fermented beans, disease resistance to witches’ broom
and abscission of mature fruits. By comparing genomic sequences and analyzing viable inter-specific hybrids, we hope to identify
the key genes that regulate cacao’s most important traits. We have investigated the utility in T. grandiflorum of three types of markers (microsatellite markers, single-strand conformational polymorphism markers and single nucleotide
polymorphism (SNP) markers) developed in cacao. Through sequencing of amplicons of 12 diverse individuals of both cacao and
T. grandiflorum, we have identified new intra- and inter-specific SNPs. Two markers which had no overlap of alleles between the species were
used to genotype putative inter-specific hybrid seedlings. Sequence conservation was significant and species-specific differences
numerous enough to suggest that comparative genomics of T. grandiflorum and T. cacao will be useful in elucidating the genetic differences that lead to a variety of important agronomic trait differences. 相似文献
2.
Current DNA extraction protocols for genomic DNA from Botrytis cinerea almost always start with mycelium that has been reduced to powder with liquid N2 in a mortar, and this makes their application to a large number of samples slow and cumbersome. Here we present an adaptation of an existing method [Möller et al. (1992) Nucleic Acids Res 20: 6115–6116] for which the initial steps have been modified, including the homogenization of the fungus with sand and the aid of a common household drill. This method allows the processing of large number of samples in much shorter times and generates an average of 4 μg DNA per sample, of sufficient quality for use in PCR and Southern blotting. 相似文献
3.
Selection of international molecular standards for DNA fingerprinting of <Emphasis Type="Italic">Theobroma cacao</Emphasis> 总被引:1,自引:0,他引:1
Saunders JA Mischke S Leamy EA Hemeida AA 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2004,110(1):41-47
A collaborative international program was initiated to identify and describe the genetic diversity of living germplasm collections of Theobroma cacao genotypes that are maintained in several international collections scattered throughout tropical cacao growing countries of the world. Simple sequence repeat (SSR) DNA analysis was identified as the most appropriate molecular tool for DNA fingerprinting these collections during an international forum representing academic, government and industry scientists in the cacao community. Twenty-five SSR primers, which had been previously described, were evaluated as potential candidates to define an efficient, standardized, molecular fingerprinting protocol for T. cacao accessions. These primers have been evaluated for reliability, widespread distribution across the cacao genome, number of alleles produced by the SSR primers in cacao and their ability to discriminate between cacao accessions. Approximately 690 cacao accessions were used to evaluate the utility of these SSR primers as international molecular standards, and a small number of test samples of T. cacao were sent to two other independent laboratories for verification. DNA fragments were selectively amplified by PCR, using the SSR primers labeled with fluorescent dyes, and separated by capillary electrophoresis. Based on this study, the 15 SSR primers that had the highest reproducibility and consistency within a common genotype, while allowing the differentiation of separate divergent genotypes, were selected as international molecular standards for DNA fingerprinting of T. cacao. 相似文献
4.
5.
Summary A micropropagation protocol was developed using cacao somatic embryo-derived plant as a source for nodal and apical stem explants,
and apical microcuttings. Microcuttings were efficiently rooted and developed into plantlets. Axillary meristems within the
remaining decapitated plantlets subsequently developed and were used for production of additional microcuttings, with an average
2.4 growing shoots per decapitated stem. The remaining plantelts were maintained as microcutting stock plants. When nodal
stem explants were cultured on thidiazuron medium, axillary buds proliferated and developed into shoots, which were excised
and rooted. However, the efficiency of this method is lower than rooting of apical microcuttings harvested directly from stock
plants. During root induction, short treatment with indole-3-butyric acid (IBA) increased the total percentage of rooted microcuttings
up to 89%. Longer exposures to IBA increased the average number of roots per microcutting (from 1.7 to 5.2). Plant acclimatization
after rooting was achieved with an average success of 87%. During several months of growth in the greenhouse, the micropropagated
plants developed functional taproots. Currently, cocoa plants produced by this micropropagation method have been successfully
acclimated to field conditions in Ivory Coast, Ghana, and Saint Lucia. 相似文献
6.
7.
Summary The development of efficient tissue culture systems for cacao holds the potential to contribute to the improvement of this
tropical erop by providing a rapid and efficient vegetative propagation system for multiplication of elite genotypes. It may
also find application in facilitation of germplasm movement across quarantine borders, enhancement of germplasm conservation
via cryo-preservation, and development of genetic transformation systems. Somatic embryogenesis using floral tissue explants
was previously the only tissue culture procedure for regeneration of cacao. We report the development of a secondary embryogenesis
system utilizing primary somatic embryo cotyledon explants, which results in up to a 30-fold increase in somatic embryo production
compared to primary somatic embryogenesis. The influence of genotype on the efficiency of the system was evaluated. To understand
the cellular origins and developmental pathways operative in this system, we investigated the morphological changes occurring
over time using light and scanning electron microscopy. While primary embryos arise from clusters of cells forming embryonic
nodules, secondary embryos arise predominantly from the division of single cells, in a pathway reminiscent of zygotic embryogenesis.
These results have important significance to the application of tissue culture to cacao improvement programs. 相似文献
8.
We describe a simple and efficient method for genomic DNA extraction from woody fruit crops containing high polysaccharide
levels. This method involves a modified CTAB or SDS procedure employing a purification step to remove polysaccharides by using
water-saturated ether and 1.25 M NaCl. Precipitation with an equal volume of isopropanol caused a DNA pellet to form. After
being washed with 70% ethyl alcohol, the pellet easily dissolved in TE buffer. Using this method, DNA was extracted from samples
of more than 1000Citrus spp., including young leaves, old leaves, frosted old leaves, withered old leaves, and callus lines. The average yield of
DNA ranged from 50–500 μg/g of sample. DNA was suitable for PCR and RFLP analyses and long-term storage. Recently, the procedure
was used to isolate DNA from withered old leaves of more than 20 tropical and subtropical fruit crops. 相似文献
9.
The mechanisms that reduce the viability of plant somatic embryos following cryopreservation are not known. The objective
of the present study was to evaluate the sensitivity of cocoa (Theobroma cacao L.) somatic embryos at different stages of an encapsulation–dehydration protocol using stress-related volatile hydrocarbons
as markers of injury and recovery. The plant stress hormone ethylene and volatile hydrocarbons derived from hydroxyl radicals
(methane) and lipid peroxidation (ethane) were determined using gas chromatography headspace analysis. Ethylene and methane
were the only volatiles detected, with both being produced after each step of the cryogenic protocol. Ethylene production
was significantly reduced following exposure to liquid nitrogen, but then increased in parallel with embryo recovery. In contrast,
the production of methane was cyclic during recovery, with the first cycle occurring earlier for embryos recovered from liquid
nitrogen and desiccation than those recovered from earlier steps in the protocol. These results suggest that loss of somatic
embryo viability during cryopreservation may be related to the oxidative status of the tissue, and its capacity to produce
ethylene. This study has demonstrated that headspace volatile analysis provides a robust non-destructive analytical approach
for assessing the survival and recovery of plant somatic embryos following cryopreservation. 相似文献
10.
Clément D Lanaud C Sabau X Fouet O Le Cunff L Ruiz E Risterucci AM Glaszmann JC Piffanelli P 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2004,108(8):1627-1634
We have constructed and validated the first cocoa (Theobroma cacao L.) BAC library, with the aim of developing molecular resources to study the structure and evolution of the genome of this perennial crop. This library contains 36,864 clones with an average insert size of 120 kb, representing approximately ten haploid genome equivalents. It was constructed from the genotype Scavina-6 (Sca-6), a Forastero clone highly resistant to cocoa pathogens and a parent of existing mapping populations. Validation of the BAC library was carried out with a set of 13 genetically-anchored single copy and one duplicated markers. An average of nine BAC clones per probe was identified, giving an initial experimental estimation of the genome coverage represented in the library. Screening of the library with a set of resistance gene analogues (RGAs), previously mapped in cocoa and co-localizing with QTL for resistance to Phytophthora traits, confirmed at the physical level the tight clustering of RGAs in the cocoa genome and provided the first insights into the relationships between genetic and physical distances in the cocoa genome. This library represents an available BAC resource for structural genomic studies or map-based cloning of genes corresponding to important QTLs for agronomic traits such as resistance genes to major cocoa pathogens like Phytophthora spp (palmivora and megakarya), Crinipellis perniciosa and Moniliophthora roreri.Communicated by J.W. Snape 相似文献
11.
Siela N. Maximova Ann Young Sharon Pishak Mark J. Guiltinan 《In vitro cellular & developmental biology. Plant》2008,44(6):487-493
Somatic embryogenesis is an in vitro clonal propagation method with potential to contribute to the improvement of cacao varieties. Before using this technology
for commercial production, it is essential that somatic embryogenesis-derived plants be tested in field conditions. Therefore,
we established a field test at Union Vale Estate, Saint Lucia. Thirty- to 50-yr-old trees were selected for clonal propagation
as potentially high yielding based on local farmers observations. Clonal plants were propagated in vitro from immature flowers by embryogenesis and micropropagation. Multiple plants from nine genotypes were acclimated to greenhouse
conditions then returned to Saint Lucia and planted in a field. Orthotropic rooted cuttings and locally propagated open pollinated
seedlings were also planted for a total of 214 trees. Growth data were collected every 4–6 mo. including: stem diameter, stem
height, length of the longest jorquette branch, number of jorquette branches, and dates of first flowering and fruiting. At
4.5 yr after planting in the field there were no major differences in all growth parameters among the propagation methods
evaluated with exception of the orthotropic rooted cuttings. Trees grown from seeds were slightly taller then trees propagated
by the other methods. Trees propagated as orthotropic rooted cuttings exhibited smaller average stem diameters, shorter stem
heights to the jorquette, and shorter jorquette branches. We concluded that somatic embryo-derived plants demonstrated normal
phenotypes in field conditions and have growth parameters similar to plants propagated by traditional methods. 相似文献
12.
Two repeated DNA sequences isolated from a partial genomic DNA library of Helianthus annuus, p HaS13 and p HaS211, were shown to represent portions of the int gene of a Ty3 /gypsy retroelement and of the RNase-Hgene of a Ty1 /copia retroelement, respectively. Southern blotting patterns obtained by hybridizing the two probes to BglII- or DraI-digested genomic DNA from different Helianthus species showed p HaS13 and p HaS211 were parts of dispersed repeats at least 8 and 7 kb in length, respectively, that were conserved in all species studied. Comparable hybridization patterns were obtained in all species with p HaS13. By contrast, the patterns obtained by hybridizing p HaS211 clearly differentiated annual species from perennials. The frequencies of p HaS13- and p HaS211-related sequences in different species were 4.3x10(4)-1.3x10(5) copies and 9.9x10(2)-8.1x10(3) copies per picogram of DNA, respectively. The frequency of p HaS13-related sequences varied widely within annual species, while no significant difference was observed among perennial species. Conversely, the frequency variation of p HaS211-related sequences was as large within annual species as within perennials. Sequences of both families were found to be dispersed along the length of all chromosomes in all species studied. However, Ty3 /gypsy-like sequences were localized preferentially at the centromeric regions, whereas Ty1/ copia-like sequences were less represented or absent around the centromeres and plentiful at the chromosome ends. These findings suggest that the two sequence families played a role in Helianthusgenome evolution and species divergence, evolved independently in the same genomic backgrounds and in annual or perennial species, and acquired different possible functions in the host genomes. 相似文献
13.
14.
Björn D. Heijstra Franz B. Pichler Quanfeng Liang Razel G. Blaza Susan J. Turner 《Antonie van Leeuwenhoek》2009,95(4):343-349
Extracellular DNA can play a structural role in the microbial environment. Here evidence is presented that an environmental
isolate of Acidovorax
temperans utilises extracellular DNA for intercellular and cell-surface attachment and that Type IV pili and electrostatic interactions
play a role in this interaction. Preliminary attempts to isolate and purify extracellular polysaccharides from A. temperans strain CB2 yielded significant amounts of DNA raising the question of whether this molecule was present as a structural component
in the extracellular matrix. The role of DNA in attachment was indicated by experiments in which the addition of DNase to
liquid medium inhibited the attachment of Acidovorax to glass wool. A Tn5 insertional mutant, lacking Type IV pili, was unable to initiate attachment. Addition of DNase caused rapid detachment of
bound cells, but no detachment occurred when proteinase, RNase or inactivated DNase were used. Addition of MgCl2 also caused significant detachment, supporting the possible mechanistic role of electrostatic interactions in the attachment
process. Although attachment was apparent in early to mid-log phase growth, surprisingly DNA was not detected in the culture
supernatant until late stationary phase and coincided with an appreciable loss of cell viability. This suggests that during
log-phase growth attachment is mediated by eDNA that is released in low quantities and/or is highly localised within the extracellular
matrix and also that stationary phase DNA release through widespread cell lysis may be a separate and unrelated event. 相似文献
15.
Eun Chan Yang Myung Sook Kim Paul John L. Geraldino Dinabandhu Sahoo Jong-Ahm Shin Sung Min Boo 《Journal of applied phycology》2008,20(2):161-168
The mitochondrial cytochrome c oxidase subunit I gene sequence was recently developed for DNA barcoding of red algal species.
We determined the 1245 base pairs of the gene from 27 taxa of an agar-producing species, Gracilaria vermiculophylla, and putative relatives and compared the results with rbcL data from the same species. A total of 392 positions (31.5%) were variable, 282 positions (22.6%) were parsimoniously informative,
and average sequence divergence was 13% in an ingroup. Within G. vermiculophylla, pairwise divergence of the gene was variable up to 11 bp (0.9%). Seven recognized haplotypes of cox1 tended to be geographically related. In the aligned 1386 bp of rbcL, three haplotypes were recognized. These results suggest that cox1 is a valuable molecular marker within species and will be very useful in haplotype analyses. 相似文献
16.
Marcos Ramos da Silva Didier Clément Karina Peres Gramacho Wilson Reis Monteiro Xavier Argout Claire Lanaud Uilson Lopes 《Tree Genetics & Genomes》2016,12(3):62
Sexual compatibility limits the production of cacao plantations, being an important selection criterion in breeding programs. However, the current method for characterizing compatibility, based on the frequency of flower setting after controlled pollination, is time consuming, requiring a long time to identify self-compatible individuals. The identification of molecular markers in genomic regions can be an alternative to allow early selection of self-compatible plants. The present study aimed to identify SNP markers associated with sexual compatibility in cacao, by utilizing genome-wide association (GWAS) mapping. A population of 295 individuals mostly from third-generation breeding populations, but also founder clones, was used. This population was phenotypically characterized by hand pollinating 8199 flowers and evaluating the flower retention 15 days after pollination. In addition, leaf samples of each individual were collected and DNA extracted for genotyping by sequencing, generating 5301 SNP markers after cleaning. Genome-wide association mapping analysis was performed using Synbreed, GCTA, and TASSEL softwares. Significant markers associated to incompatibility, likely in strong linkage disequilibrium, were found within a region of 196 kb, in the proximal end of chromosome 4, suggesting the existence of a major gene in that region. However, this result should be validated in a larger population, considering that only 295 trees were used here. When the SNP effects were treated as random in the estimation process, many other regions in the genome appears to be involved with sexual incompatibility in cacao. Candidate genes were found not only in the proximal end of chromosome 4 but also spread in several other regions of the genome. 相似文献
17.
Telomeric DNA-binding proteins (TBPs) are crucial components that regulate the structure and function of eukaryotic telomeres and are evolutionarily conserved. We have identified two homologues of AtTBP1 (for Arabidopsis thaliana telomeric DNA binding protein 1), designated as AtTBP2 and AtTRP2, which encode proteins that specifically bind to the telomeric DNA of this plant. These proteins show extensive homology with other known plant TBPs. The isolated C-terminal segments of these proteins were capable of sequence-specific binding to duplex telomeric plant DNA in vitro. DNA bending assays using the Arabidopsis TBPs revealed that AtTBP1 and AtTBP2 have DNA-bending abilities comparable to that of the human homologue hTRF1, and higher than those of AtTRP1 and AtTRP2. 相似文献
18.
The relationship between Sporidiobolus johnsonii and S. salmonicolor was investigated using rDNA sequence data. Two statistically well-supported clades were obtained. One clade included the
type strain of S. johnsonii and the other included the type strain of S. salmonicolor. However, some mating strains of S. salmonicolor were found in the S. johnsonii group. These strains belonged to mating type A2 and were sexually compatible with mating type A1 strains from the S. salmonicolor group. DNA–DNA reassociation values were high within each clade and moderate between the two clades. In the re-investigation
of teliospore germination, we observed that the basidia of S. salmonicolor were two-celled. In S. johnsonii, basidia were not formed and teliospore germination resulted in direct formation of yeast cells. We hypothesize that the
S. johnsonii clade is becoming genetically isolated from the S. salmonicolor group and that a speciation process is presently going on. We suspect that the observed sexual compatibility between strains
of the S. johnsonii and S. salmonicolor groups and the possible genetic flow between the two species has little biological relevance because distinct phenotypes
have been fixed in the two taxa and intermediate (hybrid) sequences for LSU and ITS rDNAs have not been detected.
An erratum to this article can be found at 相似文献
19.
Wild rice is a valuable resource for the genetic improvement of cultivated rice (Oryza sativa L., AA genome). Molecular markers are important tools for monitoring gene introgression from wild rice into cultivated rice. In this study, Simple sequence repeat (SSR) markers were used to analyze interspecific hybrids of O. sativa-O. officinalis (CC genome), the backcrossing progenies and the parent plants. Results showed that most of the SSR primers (335 out of 396, 84.6%) developed in cultivated rice successfully amplified products from DNA samples of wild rice O. officinalis. The polymorphism ratio of SSR bands between O. sativa and O. officinalis was as high as 93.9%, indicating differences between the two species with respect to SSRs. When the SSR markers were applied in the interspecific hybrids, only a portion of SSR primers amplified O. officinalis-specific bands in the F(1) hybrid (52.5%), BC(1) (52.5%), and MAALs (37.0%); a number of the bands disappeared. Of the 124 SSR loci that detected officinalis-specific bands in MAAL plants, 96 (77.4%) showed synteny between the A and C-genomes, and 20 (16.1%) showed duplication in the C-genome. Sequencing analysis revealed that indels, substitution and duplication contribute to the diversity of SSR loci between the genomes of O. sativa and O. officinalis. 相似文献
20.
Irene C Maciariello C Micheli G Theis JF Newlon CS Fabiani L 《Molecular genetics and genomics : MGG》2007,277(3):287-299
Eukaryotic chromosomal DNA replication is initiated by a highly conserved set of proteins that interact with cis-acting elements on chromosomes called replicators. Despite the conservation of replication initiation proteins, replicator
sequences show little similarity from species to species in the small number of organisms that have been examined. Examination
of replicators in other species is likely to reveal common features of replicators. We have examined a Kluyeromyces lactis replicator, KARS12, that functions as origin of DNA replication on plasmids and in the chromosome. It contains a 50-bp region with similarity
to two other K. lactis replicators, KARS101 and the pKD1 replication origin. Replacement of the 50-bp sequence with an EcoRI site completely abrogated the ability of KARS12 to support plasmid and chromosomal DNA replication origin activity, demonstrating this sequence is a common feature of K. lactis replicators and is essential for function, possibly as the initiator protein binding site. Additional sequences up to 1 kb
in length are required for efficient KARS12 function. Within these sequences are a binding site for a global regulator, Abf1p, and a region of bent DNA, both of which
contribute to the activity of KARS12. These elements may facilitate protein binding, protein/protein interaction and/or nucleosome positioning as has been proposed
for other eukaryotic origins of DNA replication. 相似文献