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Fulltext Mapping quantitative trait loci (QTLs) for fatty acid composition in an interspecific cross of oil palm

Singh R, Tan SG, Panandam JM, Rahman RA, Ooi LC, Low ET, et al.

BMC Plant Biol, 2009;9:114.
PMID: 19706196 DOI: 10.1186/1471-2229-9-114

Marker Assisted Selection (MAS) is well suited to a perennial crop like oil palm, in which the economic products are not produced until several years after planting. The use of DNA markers for selection in such crops can greatly reduce the number of breeding cycles needed. With the use of DNA markers, informed decisions can be made at the nursery stage, regarding which individuals should be retained as breeding stock, which are satisfactory for agricultural production, and which should be culled. The trait associated with oil quality, measured in terms of its fatty acid composition, is an important agronomic trait that can eventually be tracked using molecular markers. This will speed up the production of new and improved oil palm planting materials.

Matched MeSH terms: DNA, Plant/genetics
Fulltext Identification of non-ribosomal peptide synthetase in Ganoderma boninense Pat. that was expressed during the interaction with oil palm

Shokrollahi N, Ho CL, Zainudin NAIM, Wahab MABA, Wong MY

Sci Rep, 2021 Aug 11;11(1):16330.
PMID: 34381084 DOI: 10.1038/s41598-021-95549-8

Basal stem rot (BSR) of oil palm is a disastrous disease caused by a white-rot fungus Ganoderma boninense Pat. Non-ribosomal peptides (NRPs) synthesized by non-ribosomal peptide synthetases (NRPSs) are a group of secondary metabolites that act as fungal virulent factors during pathogenesis in the host. In this study, we aimed to isolate NRPS gene of G. boninense strain UPMGB001 and investigate the role of this gene during G. boninense-oil palm interaction. The isolated NRPS DNA fragment of 8322 bp was used to predict the putative peptide sequence of different domains and showed similarity with G. sinense (85%) at conserved motifs of three main NRPS domains. Phylogenetic analysis of NRPS peptide sequences demonstrated that NRPS of G. boninense belongs to the type VI siderophore family. The roots of 6-month-old oil palm seedlings were artificially inoculated for studying NRPS gene expression and disease severity in the greenhouse. The correlation between high disease severity (50%) and high expression (67-fold) of G. boninense NRPS gene at 4 months after inoculation and above indicated that this gene played a significant role in the advancement of BSR disease. Overall, these findings increase our knowledge on the gene structure of NRPS in G. boninense and its involvement in BSR pathogenesis as an effector gene.

Matched MeSH terms: DNA, Plant/genetics
Chromosomal Locations of a Non-LTR Retrotransposon, Menolird18, in Cucumis melo and Cucumis sativus, and Its Implication on Genome Evolution of Cucumis Species

Setiawan AB, Teo CH, Kikuchi S, Sassa H, Kato K, Koba T

Cytogenet Genome Res, 2020;160(9):554-564.
PMID: 33171461 DOI: 10.1159/000511119

Mobile elements are major regulators of genome evolution through their effects on genome size and chromosome structure in higher organisms. Non-long terminal repeat (non-LTR) retrotransposons, one of the subclasses of transposons, are specifically inserted into repetitive DNA sequences. While studies on the insertion of non-LTR retrotransposons into ribosomal RNA genes and other repetitive DNA sequences have been reported in the animal kingdom, studies in the plant kingdom are limited. Here, using FISH, we confirmed that Menolird18, a member of LINE (long interspersed nuclear element) in non-LTR retrotransposons and found in Cucumis melo, was inserted into ITS and ETS (internal and external transcribed spacers) regions of 18S rDNA in melon and cucumber. Beside the 18S rDNA regions, Menolird18 was also detected in all centromeric regions of melon, while it was located at pericentromeric and sub-telomeric regions in cucumber. The fact that FISH signals of Menolird18 were found in centromeric and rDNA regions of mitotic chromosomes suggests that Menolird18 is a rDNA and centromere-specific non-LTR retrotransposon in melon. Our findings are the first report on a non-LTR retrotransposon that is highly conserved in 2 different plant species, melon and cucumber. The clear distinction of chromosomal localization of Menolird18 in melon and cucumber implies that it might have been involved in the evolutionary processes of the melon (C. melo) and cucumber (C. sativus) genomes.

Matched MeSH terms: DNA, Plant/genetics
Fulltext DNA Barcoding, Phylogenetic Analysis and Secondary Structure Predictions of Nepenthes ampullaria, Nepenthes gracilis and Nepenthes rafflesiana

Saidon NA, Wagiran A, Samad AFA, Mohd Salleh F, Mohamed F, Jani J, et al.

Genes (Basel), 2023 Mar 11;14(3).
PMID: 36980969 DOI: 10.3390/genes14030697

Nepentheceae, the most prominent carnivorous family in the Caryophyllales order, comprises the Nepenthes genus, which has modified leaf trap characteristics. Although most Nepenthes species have unique morphologies, their vegetative stages are identical, making identification based on morphology difficult. DNA barcoding is seen as a potential tool for plant identification, with small DNA segments amplified for species identification. In this study, three barcode loci; ribulose-bisphosphate carboxylase (rbcL), intergenic spacer 1 (ITS1) and intergenic spacer 2 (ITS2) and the usefulness of the ITS1 and ITS2 secondary structure for the molecular identification of Nepenthes species were investigated. An analysis of barcodes was conducted using BLASTn, pairwise genetic distance and diversity, followed by secondary structure prediction. The findings reveal that PCR and sequencing were both 100% successful. The present study showed the successful amplification of all targeted DNA barcodes at different sizes. Among the three barcodes, rbcL was the least efficient as a DNA barcode compared to ITS1 and ITS2. The ITS1 nucleotide analysis revealed that the ITS1 barcode had more variations compared to ITS2. The mean genetic distance (K2P) between them was higher for interspecies compared to intraspecies. The results showed that the DNA barcoding gap existed among Nepenthes species, and differences in the secondary structure distinguish the Nepenthes. The secondary structure generated in this study was found to successfully discriminate between the Nepenthes species, leading to enhanced resolutions.

Matched MeSH terms: DNA, Plant/genetics
Fulltext Ser-653-Asn substitution in the acetohydroxyacid synthase gene confers resistance in weedy rice to imidazolinone herbicides in Malaysia

Ruzmi R, Ahmad-Hamdani MS, Mazlan N

PLoS One, 2020;15(9):e0227397.
PMID: 32925921 DOI: 10.1371/journal.pone.0227397

The continuous and sole dependence on imidazolinone (IMI) herbicides for weedy rice control has led to the evolution of herbicide resistance in weedy rice populations across various countries growing IMI herbicide-resistant rice (IMI-rice), including Malaysia. A comprehensive study was conducted to elucidate occurrence, level, and mechanisms endowing resistance to IMI herbicides in putative resistant (R) weedy rice populations collected from three local Malaysian IMI-rice fields. Seed bioassay and whole-plant dose-response experiments were conducted using commercial IMI herbicides. Based on the resistance index (RI) quantification in both experiments, the cross-resistance pattern of R and susceptible (S) weedy rice populations and control rice varieties (IMI-rice variety MR220CL2 and non-IMI-rice variety MR219) to imazapic and imazapyr was determined. A molecular investigation was carried out by comparing the acetohydroxyacid synthase (AHAS) gene sequences of the R and S populations and the MR220CL2 and MR219 varieties. The AHAS gene sequences of R weedy rice were identical to those of MR220CL2, exhibiting a Ser-653-Asn substitution, which was absent in MR219 and S plants. In vitro assays were conducted using analytical grade IMI herbicides of imazapic (99.3%) and imazapyr (99.6%) at seven different concentrations. The results demonstrated that the AHAS enzyme extracted from the R populations and MR220CL2 was less sensitive to IMI herbicides than that from S and MR219, further supporting that IMI herbicide resistance was conferred by target-site mutation. In conclusion, IMI resistance in the selected populations of Malaysian weedy rice could be attributed to a Ser-653-Asn mutation that reduced the sensitivity of the target site to IMI herbicides. To our knowledge, this study is the first to show the resistance mechanism in weedy rice from Malaysian rice fields.

Matched MeSH terms: DNA, Plant/genetics; DNA, Plant/isolation & purification
Biolistic-mediated production of transgenic oil palm

Parveez GK, Bahariah B

Methods Mol Biol, 2012;847:163-75.
PMID: 22351007 DOI: 10.1007/978-1-61779-558-9_14

The effectiveness of mannose (using phosphomannose isomerase [pmi] gene) as a positive selection agent to preferably allow the growth of transformed oil palm embryogenic calli was successfully evaluated. Using the above selection agent in combination with the previously optimized physical and biological parameters and the best constitutive promoter, oil palm embryogenic calli were transformed with pmi gene for producing transgenic plants. Bombarded embryogenic calli were exposed to embryogenic calli medium containing 30:0 g/L mannose to sucrose 3 weeks postbombardment. Selectively, proliferating embryogenic calli started to emerge around 6 months on the above selection medium. The proliferated embryogenic calli were individually isolated once they reached a specific size and regenerated to produce complete plantlets. The complete regenerated plantlets were evaluated for the presence of transgenes by PCR and Southern analyses.

Matched MeSH terms: DNA, Plant
Fulltext Diet Composition of the Wild Stump-Tailed Macaque (Macaca arctoides) in Perlis State Park, Peninsular Malaysia, Using a Chloroplast tRNL DNA Metabarcoding Approach: A Preliminary Study

Osman NA, Abdul-Latiff MAB, Mohd-Ridwan AR, Yaakop S, Nor SM, Md-Zain BM

Animals (Basel), 2020 Nov 26;10(12).
PMID: 33255964 DOI: 10.3390/ani10122215

Understanding dietary diversity is a fundamental task in the study of stump-tailed macaque, Macaca arctoides in its natural habitat. However, direct feeding observation and morphological identification using fecal samples are not effective and nearly impossible to obtain in natural habitats because this species is sensitive to human presence. As ecological methods are challenging and time-consuming, DNA metabarcoding offers a more powerful assessment of the diet. We used a chloroplast tRNL DNA metabarcoding approach to identify the diversity of plants consumed by free-ranging M. arctoides in the Malaysia-Thailand border region located in Perlis State Park, Peninsular Malaysia. DNA was extracted from three fecal samples, and chloroplast tRNL DNA was amplified and sequenced using the Illumina MiniSeq platform. Sequences were analyzed using the CLC Genomic Workbench software. A total of 145 plant species from 46 families were successfully identified as being consumed by M. arctoides. The most abundant species were yellow saraca, Saraca thaipingensis (11.70%), common fig, Ficus carica (9.33%), aramata, Clathrotropis brachypetala (5.90%), sea fig, Ficus superba (5.44%), and envireira, Malmea dielsiana (1.70%). However, Clathrotropis and Malmea are not considered Malaysian trees because of limited data available from Malaysian plant DNA. Our study is the first to identify plant taxa up to the species level consumed by stump-tailed macaques based on a DNA metabarcoding approach. This result provides an important understanding on diet of wild M. arctoides that only reside in Perlis State Park, Malaysia.

Matched MeSH terms: DNA, Plant
Mating system and seed variation of Acacia hybrid (A. mangium x A. auriculiformis)

Ng CH, Lee SL, Ng KK, Muhammad N, Ratnam W

J Genet, 2009 Apr;88(1):25-31.
PMID: 19417541

The mating system and seed variation of Acacia hybrid (A. mangium x A. auriculiformis) were studied using allozymes and random amplified polymorphic DNA (RAPD) markers, respectively. Multi-locus outcrossing rate estimations indicated that the hybrid was predominantly outcrossed (mean+/- s.e. t(m) = 0.86+/-0.01). Seed variation was investigated using 35 polymorphic RAPD fragments. An analysis of molecular variance (AMOVA) revealed the highest genetic variation among seeds within a pod (66%-70%), followed by among pods within inflorescence (29%-37%), and the least variation among inflorescences within tree (1%). In addition, two to four RAPD profiles could be detected among seeds within pod. Therefore, the results suggest that a maximum of four seeds per pod could be sampled for the establishment of a mapping population for further studies.

Matched MeSH terms: DNA, Plant/chemistry
A phylogenomic analysis of Nepenthes (Nepenthaceae)

Murphy B, Forest F, Barraclough T, Rosindell J, Bellot S, Cowan R, et al.

Mol Phylogenet Evol, 2020 03;144:106668.
PMID: 31682924 DOI: 10.1016/j.ympev.2019.106668

Nepenthaceae is one of the largest carnivorous plant families and features ecological and morphological adaptations indicating an impressive adaptive radiation. However, investigation of evolutionary and taxonomic questions is hindered by poor phylogenetic understanding, with previous molecular studies based on limited loci and taxa. We use high-throughput sequencing with a target-capture methodology based on a 353-loci, probe set to recover sequences for 197 samples, representing 151 described or putative Nepenthes species. Phylogenetic analyses were performed using supermatrix and maximum quartet species tree approaches. Our analyses confirm five Western outlier taxa, followed by N. danseri, as successively sister to the remainder of the group. We also find mostly consistent recovery of two major Southeast Asian clades. The first contains common or widespread lowland species plus a Wallacean-New Guinean clade. Within the second clade, sects. Insignes and Tentaculatae are well supported, while geographically defined clades representing Sumatra, Indochina, Peninsular Malaysia, Palawan, Mindanao and Borneo are also consistently recovered. However, we find considerable conflicting signal at the site and locus level, and often unstable backbone relationships. A handful of Bornean taxa are inconsistently placed and require further investigation. We make further suggestions for a modified infra-generic classification of genus Nepenthes.

Matched MeSH terms: DNA, Plant/analysis
Fulltext Scanning the effects of ethyl methanesulfonate on the whole genome of Lotus japonicus using second-generation sequencing analysis

Mohd-Yusoff NF, Ruperao P, Tomoyoshi NE, Edwards D, Gresshoff PM, Biswas B, et al.

G3 (Bethesda), 2015 Apr;5(4):559-67.
PMID: 25660167 DOI: 10.1534/g3.114.014571

Genetic structure can be altered by chemical mutagenesis, which is a common method applied in molecular biology and genetics. Second-generation sequencing provides a platform to reveal base alterations occurring in the whole genome due to mutagenesis. A model legume, Lotus japonicus ecotype Miyakojima, was chemically mutated with alkylating ethyl methanesulfonate (EMS) for the scanning of DNA lesions throughout the genome. Using second-generation sequencing, two individually mutated third-generation progeny (M3, named AM and AS) were sequenced and analyzed to identify single nucleotide polymorphisms and reveal the effects of EMS on nucleotide sequences in these mutant genomes. Single-nucleotide polymorphisms were found in every 208 kb (AS) and 202 kb (AM) with a bias mutation of G/C-to-A/T changes at low percentage. Most mutations were intergenic. The mutation spectrum of the genomes was comparable in their individual chromosomes; however, each mutated genome has unique alterations, which are useful to identify causal mutations for their phenotypic changes. The data obtained demonstrate that whole genomic sequencing is applicable as a high-throughput tool to investigate genomic changes due to mutagenesis. The identification of these single-point mutations will facilitate the identification of phenotypically causative mutations in EMS-mutated germplasm.

Matched MeSH terms: DNA, Plant/analysis; DNA, Plant/chemistry
Fulltext Genomic DNA extraction from medicinal plants available in Malaysia using a TriOmic(TM) improved extraction kit

Mohd-Hairul AR, Sade AB, Yiap BC, Raha AR

Genet. Mol. Res., 2011;10(4):2757-64.
PMID: 22095601 DOI: 10.4238/2011.November.8.1

DNA extraction was carried out on 32 medicinal plant samples available in Malaysia using the TriOmic(TM) extraction kit. Amounts of 0.1 g flowers or young leaves were ground with liquid nitrogen, lysed at 65°C in RY1(plus) buffer and followed by RNAse treatment. Then, RY2 buffer was added to the samples and mixed completely by vortexing before removal of cell debris by centrifugation. Supernatants were transferred to fresh microcentrifuge tubes and 0.1 volume RY3 buffer was added to each of the transferred supernatant. The mixtures were applied to spin columns followed by a centrifugation step to remove buffers and other residues. Washing step was carried out twice by applying 70% ethanol to the spin columns. Genomic DNA of the samples was recovered by applying 50 μL TE buffer to the membrane of each spin column, followed by a centrifugation step at room temperature. A modification of the TriOmic(TM) extraction procedure was carried out by adding chloroform:isoamyl alcohol (24:1) steps in the extraction procedure. The genomic DNA extracted from most of the 32 samples showed an increase of total yield when chloroform:isoamyl alcohol (24:1) steps were applied in the TriOmicTM extraction procedure. This preliminary study is very important for molecular studies of medicinal plants available in Malaysia since the DNA extraction can be completed in a shorter period of time (within 1 h) compared to manual extraction, which entails applying phenol, chloroform and ethanol precipitation, and requires 1-2 days to complete.

Matched MeSH terms: DNA, Plant/isolation & purification*; DNA, Plant/chemistry
Fulltext Evolution of mycoheterotrophy in Polygalaceae: The case of Epirixanthes

Mennes CB, Moerland MS, Rath M, Smets EF, Merckx VS

Am J Bot, 2015 Apr;102(4):598-608.
PMID: 25878092 DOI: 10.3732/ajb.1400549

The mycoheterotrophic lifestyle has enabled some plant lineages to obtain carbon from their mycorrhizal symbionts. The mycoheterotrophic genus Epirixanthes (Polygalaceae) consists of six species from tropical Asia. Although it is probably closely related to the chlorophyllous genus Salomonia and linked to arbuscular mycorrhizal fungi, lack of DNA sequence data has thus far prevented these hypotheses from being tested. Therefore, the evolutionary history of Epirixanthes remains largely unknown.

Matched MeSH terms: DNA, Plant/genetics
Isolation and characterization of an oil palm constitutive promoter derived from a translationally control tumor protein (TCTP) gene

Masura SS, Parveez GK, Ti LL

Plant Physiol Biochem, 2011 Jul;49(7):701-8.
PMID: 21549610 DOI: 10.1016/j.plaphy.2011.04.003

We have characterized an oil palm (Elaeis guineensis Jacq.) constitutive promoter that is derived from a translationally control tumor protein (TCTP) gene. The TCTP promoter was fused transcriptionally with the gusA reporter gene and transferred to monocot and dicot systems in order to study its regulatory role in a transient expression study. It was found that the 5' region of TCTP was capable of driving the gusA expression in all the oil palm tissues tested, including immature embryo, embryogenic callus, embryoid, young leaflet from mature palm, green leaf, mesocarp and stem. It could also be used in dicot systems as it was also capable of driving gusA expression in tobacco leaves. The results indicate that the TCTP promoter could be used for the production of recombinant proteins that require constitutive expression in the plant system.

Matched MeSH terms: DNA, Plant/genetics; DNA, Plant/chemistry
Fulltext Non-density dependent pollen dispersal of Shorea maxwelliana (Dipterocarpaceae) revealed by a Bayesian mating model based on paternity analysis in two synchronized flowering seasons

Masuda S, Tani N, Ueno S, Lee SL, Muhammad N, Kondo T, et al.

PLoS One, 2013;8(12):e82039.
PMID: 24391712 DOI: 10.1371/journal.pone.0082039

Pollinator syndrome is one of the most important determinants regulating pollen dispersal in tropical tree species. It has been widely accepted that the reproduction of tropical forest species, especially dipterocarps that rely on insects with weak flight for their pollination, is positively density-dependent. However differences in pollinator syndrome should affect pollen dispersal patterns and, consequently, influence genetic diversity via the mating process. We examined the pollen dispersal pattern and mating system of Shorea maxwelliana, the flowers of which are larger than those of Shorea species belonging to section Mutica which are thought to be pollinated by thrips (weak flyers). A Bayesian mating model based on the paternity of seeds collected from mother trees during sporadic and mass flowering events revealed that the estimated pollen dispersal kernel and average pollen dispersal distance were similar for both flowering events. This evidence suggests that the putative pollinators - small beetles and weevils - effectively contribute to pollen dispersal and help to maintain a high outcrossing rate even during sporadic flowering events. However, the reduction in pollen donors during a sporadic event results in a reduction in effective pollen donors, which should lead to lower genetic diversity in the next generation derived from seeds produced during such an event. Although sporadic flowering has been considered less effective for outcrossing in Shorea species that depend on thrips for their pollination, effective pollen dispersal by the small beetles and weevils ensures outcrossing during periods of low flowering tree density, as occurs in a sporadic flowering event.

Matched MeSH terms: DNA, Plant/genetics
Fulltext Pollination implications of the diverse diet of tropical nectar-feeding bats roosting in an urban cave

Lim VC, Ramli R, Bhassu S, Wilson JJ

PeerJ, 2018;6:e4572.
PMID: 29607265 DOI: 10.7717/peerj.4572

Background: Intense landscaping often alters the plant composition in urban areas. Knowing which plant species that pollinators are visiting in urban areas is necessary for understanding how landscaping impacts biodiversity and associated ecosystem services. The cave nectar bat,Eonycteris spelaea, is an important pollinator for many plants and is often recorded in human-dominated habitats. Previous studies of the diet ofE. spelaearelied on morphological identification of pollen grains found in faeces and on the body of bats and by necessity disregarded other forms of digested plant material present in the faeces (i.e., plant juice and remnants). The main objective of this study was to examine the diet of the nectarivorous bat,E. spelaea,roosting in an urban cave at Batu Caves, Peninsular Malaysia by identifying the plant material present in the faeces of bats using DNA metabarcoding.
Methods: Faeces were collected under the roost ofE. spelaeaonce a week from December 2015 to March 2016. Plant DNA was extracted from the faeces, Polymerase chain reaction (PCR) amplified atITS2andrbcLregions and mass sequenced. The resultant plant operational taxonomic units were searched against NCBI GenBank for identification.
Results: A total of 55 species of plants were detected from faeces ofE. spelaeaincludingArtocarpus heterophyllus, Duabanga grandifloraandMusaspp. which are likely to be important food resources for the cave nectar bat.
Discussion: Many native plant species that had not been reported in previous dietary studies ofE. spelaeawere detected in this study includingBauhinia strychnoideaandUrophyllum leucophlaeum, suggesting thatE. spelaearemains a crucial pollinator for these plants even in highly disturbed habitats. The detection of many introduced plant species in the bat faeces indicates thatE. spelaeaare exploiting them, particularlyXanthostemon chrysanthus,as food resources in urban area. Commercial food crops were detected from all of the faecal samples, suggesting thatE. spelaeafeed predominantly on the crops particularly jackfruit and banana and play a significant role in pollination of economically important plants. Ferns and figs were also detected in the faeces ofE. spelaeasuggesting future research avenues to determine whether the 'specialised nectarivorous'E. spelaeafeed opportunistically on other parts of plants.

Matched MeSH terms: DNA, Plant
Fulltext Epigenetic Distribution of Recombinant Plant Chromosome Fragments in a Human-Arabidopsis Hybrid Cell Line

Liaw Y, Liu Y, Teo C, Cápal P, Wada N, Fukui K, et al.

Int J Mol Sci, 2021 May 21;22(11).
PMID: 34063996 DOI: 10.3390/ijms22115426

Methylation systems have been conserved during the divergence of plants and animals, although they are regulated by different pathways and enzymes. However, studies on the interactions of the epigenomes among evolutionarily distant organisms are lacking. To address this, we studied the epigenetic modification and gene expression of plant chromosome fragments (~30 Mb) in a human-Arabidopsis hybrid cell line. The whole-genome bisulfite sequencing results demonstrated that recombinant Arabidopsis DNA could retain its plant CG methylation levels even without functional plant methyltransferases, indicating that plant DNA methylation states can be maintained even in a different genomic background. The differential methylation analysis showed that the Arabidopsis DNA was undermethylated in the centromeric region and repetitive elements. Several Arabidopsis genes were still expressed, whereas the expression patterns were not related to the gene function. We concluded that the plant DNA did not maintain the original plant epigenomic landscapes and was under the control of the human genome. This study showed how two diverging genomes can coexist and provided insights into epigenetic modifications and their impact on the regulation of gene expressions between plant and animal genomes.

Matched MeSH terms: DNA, Plant/genetics
Fulltext Satellite DNA in Paphiopedilum subgenus Parvisepalum as revealed by high-throughput sequencing and fluorescent in situ hybridization

Lee YI, Yap JW, Izan S, Leitch IJ, Fay MF, Lee YC, et al.

BMC Genomics, 2018 Aug 02;19(1):578.
PMID: 30068293 DOI: 10.1186/s12864-018-4956-7

BACKGROUND: Satellite DNA is a rapidly diverging, largely repetitive DNA component of many eukaryotic genomes. Here we analyse the evolutionary dynamics of a satellite DNA repeat in the genomes of a group of Asian subtropical lady slipper orchids (Paphiopedilum subgenus Parvisepalum and representative species in the other subgenera/sections across the genus). A new satellite repeat in Paphiopedilum subgenus Parvisepalum, SatA, was identified and characterized using the RepeatExplorer pipeline in HiSeq Illumina reads from P. armeniacum (2n = 26). Reconstructed monomers were used to design a satellite-specific fluorescent in situ hybridization (FISH) probe. The data were also analysed within a phylogenetic framework built using the internal transcribed spacer (ITS) sequences of 45S nuclear ribosomal DNA.
RESULTS: SatA comprises c. 14.5% of the P. armeniacum genome and is specific to subgenus Parvisepalum. It is composed of four primary monomers that range from 230 to 359 bp and contains multiple inverted repeat regions with hairpin-loop motifs. A new karyotype of P. vietnamense (2n = 28) is presented and shows that the chromosome number in subgenus Parvisepalum is not conserved at 2n = 26, as previously reported. The physical locations of SatA sequences were visualised on the chromosomes of all seven Paphiopedilum species of subgenus Parvisepalum (2n = 26-28), together with the 5S and 45S rDNA loci using FISH. The SatA repeats were predominantly localisedin the centromeric, peri-centromeric and sub-telocentric chromosome regions, but the exact distribution pattern was species-specific.
CONCLUSIONS: We conclude that the newly discovered, highly abundant and rapidly evolving satellite sequence SatA is specific to Paphiopedilum subgenus Parvisepalum. SatA and rDNA chromosomal distributions are characteristic of species, and comparisons between species reveal that the distribution patterns generate a strong phylogenetic signal. We also conclude that the ancestral chromosome number of subgenus Parvisepalum and indeed of all Paphiopedilum could be either 2n = 26 or 28, if P. vietnamense is sister to all species in the subgenus as suggested by the ITS data.

Matched MeSH terms: DNA, Plant/genetics
Fulltext DNA Barcoding of the Endangered Aquilaria (Thymelaeaceae) and Its Application in Species Authentication of Agarwood Products Traded in the Market

Lee SY, Ng WL, Mahat MN, Nazre M, Mohamed R

PLoS One, 2016;11(4):e0154631.
PMID: 27128309 DOI: 10.1371/journal.pone.0154631

The identification of Aquilaria species from their resinous non-wood product, the agarwood, is challenging as conventional techniques alone are unable to ascertain the species origin. Aquilaria is a highly protected species due to the excessive exploitation of its precious agarwood. Here, we applied the DNA barcoding technique to generate barcode sequences for Aquilaria species and later applied the barcodes to identify the source species of agarwood found in the market. We developed a reference DNA barcode library using eight candidate barcode loci (matK, rbcL, rpoB, rpoC1, psbA-trnH, trnL-trnF, ITS, and ITS2) amplified from 24 leaf accessions of seven Aquilaria species obtained from living trees. Our results indicated that all single barcodes can be easily amplified and sequenced with the selected primers. The combination of trnL-trnF+ITS and trnL-trnF+ITS2 yielded the greatest species resolution using the least number of loci combination, while matK+trnL-trnF+ITS showed potential in detecting the geographical origins of Aquilaria species. We propose trnL-trnF+ITS2 as the best candidate barcode for Aquilaria as ITS2 has a shorter sequence length compared to ITS, which eases PCR amplification especially when using degraded DNA samples such as those extracted from processed agarwood products. A blind test conducted on eight agarwood samples in different forms using the proposed barcode combination proved successful in their identification up to the species level. Such potential of DNA barcoding in identifying the source species of agarwood will contribute to the international timber trade control, by providing an effective method for species identification and product authentication.

Matched MeSH terms: DNA, Plant/genetics*
Inheritance studies of SSR and ISSR molecular markers and phylogenetic relationship of rice genotypes resistant to tungro virus

Latif MA, Rahman MM, Ali ME, Ashkani S, Rafii MY

C. R. Biol., 2013 Mar;336(3):125-33.
PMID: 23643394 DOI: 10.1016/j.crvi.2012.12.002

Multivariate analyses were performed using 13 morphological traits and 13 molecular markers (10 SSRs and three ISSRs) to assess the phylogenetic relationship among tungro resistant genotypes. For morphological traits, the genotypes were grouped into six clusters, according to D(2) statistic and Canonical vector analysis. Plant height, days to flowering, days to maturity, panicle length, number of spikelet per panicle, number of unfilled grain per panicle and yield were important contributors to genetic divergence in 14 rice genotypes. Based on Nei's genetic distance for molecular studies, seven clusters were formed among the tungro resistant and susceptible genotypes. Mantel's test revealed a significant correlation (r = 0.834*) between the morphological and molecular data. To develop high yielding tungro resistant varieties based on both morphological and molecular analyses, crosses could be made with susceptible (BR10 and BR11) genotypes with low yielding but highly resistant genotypes, Sonahidemota, Kumragoir, Nakuchimota, Khaiyamota, Khairymota and Kachamota. The chi-square analysis for seven alleles (RM11, RM17, RM20, RM23, RM80, RM108 and RM531) of SSR and five loci (RY1, MR1, MR2, MR4 and GF5) of three ISSR markers in F2 population of cross, BR11×Sonahidemota, showed a good fit to the expected segregation ratio (1:2:1) for a single gene model.

Matched MeSH terms: DNA, Plant/genetics
Fulltext Food assimilated by two sympatric populations of the brown planthopper Nilaparvata lugens (Delphacidae) feeding on different host plants contaminates insect DNA detected by RAPD-PCR analysis

Latif MA, Omar MY, Tan SG, Siraj SS, Ali ME, Rafii MY

Genet. Mol. Res., 2012;11(1):30-41.
PMID: 22290463 DOI: 10.4238/2012.January.9.4

Contamination of insect DNA for RAPD-PCR analysis can be a problem because many primers are non-specific and DNA from parasites or gut contents may be simultaneously extracted along with that of the insect. We measured the quantity of food ingested and assimilated by two sympatric populations of brown planthopper (BPH), Nilaparvata lugens, one from rice and the other from Leersia hexandra (Poaceae), a wetland forage grass, and we also investigated whether host plant DNA contaminates that of herbivore insects in extractions of whole insects. Ingestion and assimilation of food were reduced significantly when individuals derived from one host plant were caged on the other species. The bands, OPA3 (1.25), OPD3 (1.10), OPD3 (0.80), OPD3 (0.60), pUC/M13F (0.35), pUC/M13F (0.20), BOXAIR (0.50), peh#3 (0.50), and peh#3 (0.17) were found in both rice-infesting populations of brown planthopper and its host plant (rice). Similarly, the bands, OPA4 (1.00), OPB10 (0.70), OPD3 (0.90), OPD3 (0.80), OPD3 (0.60), pUC/ M13F (0.35), pUC/M13F (0.20), and BOXAIR (0.50) were found in both Leersia-infesting populations of brown planthopper and the host plant. So, it is clear that the DNA bands amplified in the host plants were also found in the extracts from the insects feeding on them.

Matched MeSH terms: DNA, Plant/genetics*

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