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1.
Front Plant Sci ; 15: 1347744, 2024.
Article in English | MEDLINE | ID: mdl-38504891

ABSTRACT

Introduction: Lablab (Lablab purpureus (L.) Sweet), an underutilized tropical legume crop, plays a crucial role in global food and nutritional security. To enhance our understanding of its genetic makeup towards developing elite cultivars, we sequenced and assembled a draft genome of L. purpureus accession PK2022T020 using a single tube long fragment read (stLFR) technique. Results and discussion: The preliminary assembly encompassed 367 Mb with a scaffold N50 of 4.3 Mb. To improve the contiguity of our draft genome, we employed a chromatin contact mapping (Hi-C) approach to obtain a pseudochromosome-level assembly containing 366 Mb with an N50 length of 31.1 Mb. A total of 327.4 Mb had successfully been anchored into 11 pseudomolecules, corresponding to the haploid chromosome number in lablab. Our gene prediction recovered 98.4% of the highly conserved orthologs based on the Benchmarking Universal Single-Copy Orthologs (BUSCO) analysis. Comparative analyses utilizing sequence information from single-copy orthologous genes demonstrated that L. purpureus diverged from the last common ancestor of the Phaseolus/Vigna species approximately 27.7 million years ago. A gene family expansion analysis revealed a significant expansion of genes involved in responses to biotic and abiotic stresses. Our high-quality chromosome-scale reference assembly provides an invaluable genomic resource for lablab genetic improvement and future comparative genomics studies among legume species.

2.
Mitochondrial DNA B Resour ; 8(9): 932-935, 2023.
Article in English | MEDLINE | ID: mdl-37674910

ABSTRACT

Heritiera fomes Buch.-Ham. (1800) is a species of mangrove in the family Malvaceae, widely distributed in the Indo-Pacific and listed as 'endangered' (EN) on the International Union for Conservation of Nature's (IUCN) red list. We reported the complete chloroplast genome sequence of H. fomes. The genome was 168,521 bp in length and included two inverted repeats (IRs) of 34,496 bp, separated by a large single-copy (LSC) region of 88,604 bp and a small single-copy (SSC) region of 10,925 bp, respectively. The genome contained 87 protein-coding genes (PCGs), 8 rRNA genes, and 37 tRNA genes. The maximum-likelihood (ML) phylogenetic tree suggested that H. fomes is closely related to Heritiera angustata and Heritiera parvifolia with relatively high support bootstrap values of 86% and 100% with other species (Heritiera littoralis and Heritiera javanica), suggesting a relatively close genetic relationship between the five Heritiera plants. The chloroplast genome sequence provided a useful resource for conservation genetics studies of H. fomes and for phylogenetic studies of Heritiera.

3.
Sci Rep ; 12(1): 20474, 2022 11 28.
Article in English | MEDLINE | ID: mdl-36443360

ABSTRACT

Sugarcane accounts for a large portion of the worlds sugar production. Modern commercial cultivars are complex hybrids of S. officinarum, S. spontaneum, and several other Saccharum species, resulting in an auto-allopolyploid with 8-12 copies of each chromosome. The current genome assembly gold standard is to generate a long read assembly followed by chromatin conformation capture sequencing to scaffold. We used the PacBio RSII and chromatin conformation capture sequencing to sequence and assemble the genome of a South East Asian commercial sugarcane cultivar, known as Khon Kaen 3. The Khon Kaen 3 genome assembled into 104,477 contigs totalling 7 Gb, which scaffolded into 56 pseudochromosomes containing 5.2 Gb of sequence. Genome annotation produced 242,406 genes from 30,927 orthogroups. Aligning the Khon Kaen 3 genome sequence to S. officinarum and S. spontaneum revealed a high level of apparent recombination, indicating a chimeric assembly. This assembly error is explained by high nucleotide identity between S. officinarum and S. spontaneum, where 91.8% of S. spontaneum aligns to S. officinarum at 94% identity. Thus, the subgenomes of commercial sugarcane are so similar that using short reads to correct long PacBio reads produced chimeric long reads. Future attempts to sequence sugarcane must take this information into account.


Subject(s)
Saccharum , Saccharum/genetics , Thailand , Chromatin , Edible Grain , Sequence Analysis, DNA
4.
DNA Res ; 29(5)2022 Aug 23.
Article in English | MEDLINE | ID: mdl-36214371

ABSTRACT

Bats (Chiroptera) constitute the second largest order of mammals and have several distinctive features, such as true self-powered flight and strong immunity. The Pendlebury's roundleaf bat, Hipposideros pendleburyi, is endemic to Thailand and listed as a vulnerable species. We employed the 10× Genomics linked-read technology to obtain a genome assembly of H. pendleburyi. The assembly size was 2.17 Gb with a scaffold N50 length of 15,398,518 bases. Our phylogenetic analysis placed H. pendleburyi within the rhinolophoid clade of the suborder Yinpterochiroptera. A synteny analysis showed that H. pendleburyi shared conserved chromosome segments (up to 105 Mb) with Rhinolophus ferrumequinum and Phyllostomus discolor albeit having different chromosome numbers and belonging different families. We found positive selection signals in genes involved in inflammation, spermatogenesis and Wnt signalling. The analyses of transposable elements suggested the contraction of short interspersed nuclear elements (SINEs) and the accumulation of young mariner DNA transposons in the analysed hipposiderids. Distinct mariners were likely horizontally transferred to hipposiderid genomes over the evolution of this family. The lineage-specific profiles of SINEs and mariners might involve in the evolution of hipposiderids and be associated with the phylogenetic separations of these bats from other bat families.


Subject(s)
Chiroptera , DNA Transposable Elements , Animals , Chiroptera/genetics , Genomics , Humans , Phylogeny
5.
Biology (Basel) ; 11(10)2022 Oct 12.
Article in English | MEDLINE | ID: mdl-36290398

ABSTRACT

Mitragyna speciosa (Kratom) is a tropical narcotic plant native to Southeast Asia with unique pharmacological properties. Here, we report the first chromosome-scale assembly of the M. speciosa genome. We employed PacBio sequencing to obtain a preliminary assembly, which was subsequently scaffolded using the chromatin contact mapping technique (Hi-C) into 22 pseudomolecules. The final assembly was 692 Mb with a scaffold N50 of 26 Mb. We annotated a total of 39,708 protein-coding genes, and our gene predictions recovered 98.4% of the highly conserved orthologs based on the BUSCO analysis. The phylogenetic analysis revealed that M. speciosa diverged from the last common ancestors of Coffea arabica and Coffea canephora approximately 47.6 million years ago. Our analysis of the sequence divergence at fourfold-degenerate sites from orthologous gene pairs provided evidence supporting a genome-wide duplication in M. speciosa, agreeing with the report that members of the genus Mitragyna are tetraploid. The STRUCTURE and principal component analyses demonstrated that the 85 M. speciosa accessions included in this study were an admixture of two subpopulations. The availability of our high-quality chromosome-level genome assembly and the transcriptomic resources will be useful for future studies on the alkaloid biosynthesis pathway, as well as comparative phylogenetic studies in Mitragyna and related species.

6.
Biology (Basel) ; 11(3)2022 Feb 28.
Article in English | MEDLINE | ID: mdl-35336757

ABSTRACT

Ceriops and Avicennia are true mangroves in the middle and seaward zones of mangrove forests, respectively. The chloroplast genomes of Ceriops decandra, Ceriops zippeliana, and Ceriops tagal were assembled into lengths of 166,650, 166,083 and 164,432 bp, respectively, whereas Avicennia lanata was 148,264 bp in length. The gene content and gene order are highly conserved among these species. The chloroplast genome contains 125 genes in A. lanata and 129 genes in Ceriops species. Three duplicate genes (rpl2, rpl23, and trnM-CAU) were found in the IR regions of the three Ceriops species, resulting in expansion of the IR regions. The rpl32 gene was lost in C. zippeliana, whereas the infA gene was present in A. lanata. Short repeats (<40 bp) and a lower number of SSRs were found in A. lanata but not in Ceriops species. The phylogenetic analysis supports that all Ceriops species are clustered in Rhizophoraceae and A. lanata is in Acanthaceae. In a search for genes under selective pressures of coastal environments, the rps7 gene was under positive selection compared with non-mangrove species. Finally, two specific primer sets were developed for species identification of the three Ceriops species. Thus, this finding provides insightful genetic information for evolutionary relationships and molecular markers in Ceriops and Avicennia species.

7.
Genome Biol Evol ; 14(2)2022 02 04.
Article in English | MEDLINE | ID: mdl-35148390

ABSTRACT

Upriver orange mangrove (Bruguiera sexangula) is a member of the most mangrove-rich taxon (Rhizophoraceae family) and is commonly distributed in the intertidal zones in tropical and subtropical latitudes. In this study, we employed the 10× Genomics linked-read technology to obtain a preliminary de novo assembly of the B. sexangula genome, which was further scaffolded to a pseudomolecule level using the Bruguiera parviflora genome as a reference. The final assembly of the B. sexangula genome contained 260 Mb with an N50 scaffold length of 11,020,310 bases. The assembly comprised 18 pseudomolecules (corresponding to the haploid chromosome number in B. sexangula), covering 204,645,832 bases or 78.6% of the 260-Mb assembly. We predicted a total of 23,978 protein-coding sequences, 17,598 of which were associated with gene ontology terms. Our gene prediction recovered 96.6% of the highly conserved orthologs based on the Benchmarking Universal Single-Copy Orthologs (BUSCO) analysis. The chromosome-level assembly presented in this work provides a valuable genetic resource to help strengthen our understanding of mangroves' physiological and morphological adaptations to the intertidal zones.


Subject(s)
Citrus sinensis , Rhizophoraceae , Chromosomes , Citrus sinensis/genetics , Genome , Genomics , Rhizophoraceae/genetics
8.
Gigascience ; 122022 12 28.
Article in English | MEDLINE | ID: mdl-37470496

ABSTRACT

Vigna reflexo-pilosa (créole bean) is a wild legume belonging to the subgenus Ceratoropis and is widely distributed in Asia. Créole bean is the only tetraploid species in the genus Vigna, and it has been shown to derive from the hybridization of Vigna hirtella and Vigna trinervia. In this study, we combined the long-read PacBio technology with the chromatin contact mapping (Hi-C) technique to obtain a chromosome-level assembly of V. reflexo-pilosa. The final assembly contained 998,724,903 bases with an N50 length of 42,545,650 bases. Our gene prediction recovered 99.4% of the highly conserved orthologs based on the BUSCO analysis. To investigate homoeolog expression bias and expression level dominance in the tetraploid, we also sequenced and assembled the genomes of its progenitors. Overall, the majority of the homoeolog pairs (72.9%) displayed no expression bias, and among those that exhibited biased expression, 16.3% showed unbalanced homoeolog expression bias toward the V. trinervia subgenome. Moreover, 41.2% and 36.2% of the expressed gene pairs exhibited transgressive expression and expression level dominance, respectively. Interestingly, the genome-wide expression level dominance in the tetraploid was biased toward the V. trinervia subgenome. The analysis of methylation patterns also revealed that the average methylation levels in coding regions were higher in the V. hirtella subgenome than those in the V. trinervia subgenome. The genomic/transcriptomic resources for these three species are useful not only for the development of elite cultivars in Vigna breeding programs but also to researchers studying comparative genomics and investigating genomic/epigenomic changes following polyploid events.


Subject(s)
Chrysobalanaceae , Fabaceae , Vigna , Vigna/genetics , Chrysobalanaceae/genetics , Tetraploidy , Plant Breeding , Fabaceae/genetics , Genome, Plant
9.
PLoS One ; 16(11): e0260468, 2021.
Article in English | MEDLINE | ID: mdl-34843573

ABSTRACT

Centella asiatica is rich in medical and cosmetic properties. While physiological responses of C. asiatica to light have been widely reported, the knowledge of the effects of light on its gene expression is sparse. In this study, we used RNA sequencing (RNA-seq) to investigate the expression of the C. asiatica genes in response to monochromatic red and blue light. Most of the differentially expressed genes (DEGs) under blue light were up-regulated but those under red light were down-regulated. The DEGs encoded for CRY-DASH and UVR3 were among up-regulated genes that play significant roles in responses under blue light. The DEGs involved in the response to photosystem II photodamages and in the biosynthesis of photoprotective xanthophylls were also up-regulated. The expression of flavonoid biosynthetic DEGs under blue light was up-regulated but that under red light was down-regulated. Correspondingly, total flavonoid content under blue light was higher than that under red light. The ABI5, MYB4, and HYH transcription factors appeared as hub nodes in the protein-protein interaction network of the DEGs under blue light while ERF38 was a hub node among the DEGs under red light. In summary, stress-responsive genes were predominantly up-regulated under blue light to respond to stresses that could be induced under high energy light. The information obtained from this study can be useful to better understand the responses of C. asiatica to different light qualities.


Subject(s)
Centella/genetics , Gene Expression Regulation, Plant/radiation effects , Transcriptome/radiation effects , Centella/radiation effects , Genes, Plant/radiation effects , Light , Stress, Physiological/radiation effects
10.
Sci Rep ; 11(1): 19624, 2021 10 04.
Article in English | MEDLINE | ID: mdl-34608174

ABSTRACT

The putative ferricrocin synthetase gene ferS in the fungal entomopathogen Beauveria bassiana BCC 2660 was identified and characterized. The 14,445-bp ferS encodes a multimodular nonribosomal siderophore synthetase tightly clustered with Fusarium graminearum ferricrocin synthetase. Functional analysis of this gene was performed by disruption with the bar cassette. ΔferS mutants were verified by Southern and PCR analyses. HPLC and TLC analyses of crude extracts indicated that biosynthesis of ferricrocin was abolished in ΔferS. Insect bioassays surprisingly indicated that ΔferS killed the Spodoptera exigua larvae faster (LT50 59 h) than wild type (66 h). Growth and developmental assays of the mutant and wild type demonstrated that ΔferS had a significant increase in germination under iron depletion and radial growth and a decrease in conidiation. Mitotracker staining showed that the mitochondrial activity was enriched in ΔferS under both iron excess and iron depletion. Comparative transcriptomes between wild type and ΔferS indicated that the mutant was increased in the expression of eight cytochrome P450 genes and those in iron homeostasis, ferroptosis, oxidative stress response, ergosterol biosynthesis, and TCA cycle, compared to wild type. Our data suggested that ΔferS sensed the iron excess and the oxidative stress and, in turn, was up-regulated in the antioxidant-related genes and those in ergosterol biosynthesis and TCA cycle. These increased biological pathways help ΔferS grow and germinate faster than the wild type and caused higher insect mortality than the wild type in the early phase of infection.


Subject(s)
Beauveria/growth & development , Beauveria/metabolism , Ferrichrome/analogs & derivatives , Host-Pathogen Interactions , Insecta/microbiology , Iron/metabolism , Animals , Beauveria/classification , Beauveria/pathogenicity , Computational Biology , Ferrichrome/metabolism , Fungal Proteins/genetics , Fungal Proteins/metabolism , Gene Expression Profiling , Gene Expression Regulation, Enzymologic , Gene Expression Regulation, Fungal , Homeostasis , Mutation , Oxidative Stress , Phylogeny , Virulence/genetics
11.
Genomics ; 113(4): 2717-2729, 2021 07.
Article in English | MEDLINE | ID: mdl-34089786

ABSTRACT

Corals live with complex assemblages of microbes including bacteria, the dinoflagellate Symbiodiniaceae, fungi and viruses in a coral holobiont. These coral-associated microorganisms play an important role in their host fitness and survival. Here, we investigated the structure and diversity of algal and bacterial communities associated with five Indo-Pacific coral species, using full-length 16S rRNA and internal transcribed spacer sequences. While the dinoflagellate communities associated with Poriteslutea were dominated with Symbiodiniaceae genus Cladocopium, the other four coral hosts were associated mainly with members of the Durusdinium genus, suggesting that host species was one of the underlying factors influencing the structure and composition of dinoflagellate communities associated with corals in the Gulf of Thailand. Alphaproteobacteria dominated the microbiomes of Pocillopora spp. while Pavonafrondifera and P. lutea were associated primarily with Gammaproteobacteria. Finally, we demonstrated a superior performance of full-length 16S rRNA sequences in achieving species-resolution taxonomic classification of coral-associated microbiota.


Subject(s)
Anthozoa , Dinoflagellida , Animals , Anthozoa/genetics , Bacteria/genetics , Dinoflagellida/genetics , Genes, rRNA , RNA, Ribosomal, 16S/genetics , Thailand
12.
Genomics ; 113(4): 2221-2228, 2021 07.
Article in English | MEDLINE | ID: mdl-34022344

ABSTRACT

Centella asiatica is a herbaceous, perennial species indigenous to India and Southeast Asia. C. asiatica possesses several medicinal properties: anti-aging, anti-inflammatory, wound healing and memory enhancing. The lack of available genomics resources significantly impedes the improvement of C. asiatica varieties through molecular breeding. Here, we combined the 10× Genomics linked-read technology and the long-range HiC technique to obtain the genome assembly. The final assembly contained nine pseudomolecules, corresponding to the haploid chromosome number in C. asiatica. These nine chromosomes covered 402,536,584 bases or 93.6% of the 430-Mb assembly. Comparative genomics analyses based on single-copy orthologous genes showed that C. asiatica and the common ancestor of Coriandrum sativum (coriander) and Daucus carota (carrot) diverged about 48 million years ago. This assembly provides a valuable reference genome for future molecular studies, varietal development through marker-assisted breeding and comparative genomics studies in C. asiatica.


Subject(s)
Centella , Centella/genetics , Chromosomes , Genome , Genomics/methods , Plant Breeding
13.
Mol Ecol Resour ; 21(1): 212-225, 2021 Jan.
Article in English | MEDLINE | ID: mdl-32841550

ABSTRACT

Luffa spp. (sponge gourd or ridge gourd) is an economically important vegetable crop widely cultivated in China, India and Southeast Asia. Here, we employed PacBio long-read single-molecule real-time (SMRT) sequencing to perform de novo genome assemblies of two commonly cultivated Luffa species, L. acutangula and L. cylindrica. We obtained preliminary draft genomes of 734.6 Mb and 689.8 Mb with scaffold N50 of 786,130 and 578,616 bases for L. acutangula and L. cylindrica, respectively. We also applied long-range Chicago and HiC techniques to obtain the first chromosome-scale whole-genome assembly of L. acutangula. The final assembly contained 13 pseudomolecules, corresponding to the haploid chromosome number in Luffa spp. (1n = 13, 2n = 26). The sizes of the assembled Luffa genomes are approximately twice as large as the genome assemblies of related Cucurbitaceae. A large proportion of L. acutangula (62.17%; 456.69 Mb) and L. cylindrica (56.78%; 391.65 Mb) genome assemblies contained repetitive elements. Phylogenetic analyses revealed that the substantial accumulation of transposable elements likely contributed to the expansion of the Luffa genomes. We also investigated alternative splicing events in Luffa using full-length transcript sequences obtained from PacBio Isoform Sequencing (Iso-seq). While the predominant form of alternative splicing in most plant species examined was intron retention, alternative 3' acceptor site selection appeared to be a major event observed in Luffa. High-quality genome assemblies for L. acutangula and L. cylindrica reported here provide valuable resources for Luffa breeding and future genetics and comparative genomics studies in Cucurbitaceae.


Subject(s)
DNA Transposable Elements , Genome, Plant , Luffa , Genome Size , Luffa/genetics , Phylogeny , Plant Breeding
14.
Mol Ecol Resour ; 21(1): 238-250, 2021 Jan.
Article in English | MEDLINE | ID: mdl-32794377

ABSTRACT

Black gram (Vigna mungo) is an important short duration grain legume crop. Black gram seeds provide an inexpensive source of dietary protein. Here, we applied the 10X Genomics linked-read technology to obtain a de novo whole genome assembly of V. mungo cultivated variety Chai Nat 80 (CN80). The preliminary assembly contained 12,228 contigs and had an N50 length of 5.2 Mb. Subsequent scaffolding using the long-range Chicago and HiC techniques yielded the first high-quality, chromosome-level assembly of 499 Mb comprising 11 pseudomolecules. Comparative genomics analyses based on sequence information from single-copy orthologous genes revealed that black gram and mungbean (Vigna radiata) diverged about 2.7 million years ago . The transversion rate (4DTv) analysis in V. mungo revealed no evidence supporting a recent genome-wide duplication event observed in the tetraploid créole bean (Vigna reflexo-pilosa). The proportion of repetitive elements in the black gram genome is slightly lower than the numbers reported for related Vigna species. The majority of long terminal repeat retrotransposons appeared to integrate into the genome within the last five million years. We also examined alternative splicing events in V. mungo using full-length transcript sequences. While intron retention was the most prevalent mode of alternative splicing in several plant species, alternative 3' acceptor site selection represented the majority of events in black gram. Our high-quality genome assembly along with the genomic variation information from the germplasm provides valuable resources for accelerating the development of elite varieties through marker-assisted breeding and for future comparative genomics and phylogenetic studies in legume species.


Subject(s)
Genome, Plant , Vigna , Chromosomes, Plant , Crops, Agricultural/genetics , Phylogeny , Plant Breeding , Retroelements , Vigna/genetics
15.
Data Brief ; 33: 106470, 2020 Dec.
Article in English | MEDLINE | ID: mdl-33195780

ABSTRACT

Luffa acutangula and Luffa aegyptiaca are domesticated plants in the family Cucurbitaceae. They are mainly cultivated in the tropical and subtropical regions of Asia. The chloroplast genomes of many Cucurbitaceae species were sequenced to examine gene content and evolution. However, the chloroplast genome sequences of L. acutangula and L. aegyptiaca have not been reported. We report the first complete sequences of L. acutangula and L. aegyptiaca chloroplast genomes obtained from Pacific Biosciences sequencing and use them to infer evolutionary relationships. The chloroplast genomes of L. acutangula and L. aegyptiaca are 157,202 and 157,275 bp, respectively. Both genomes possessed the typical quadripartite structure and contained 131 genes, including 87 coding genes, 36 tRNA genes and 8 rRNA genes. We identified simple sequence repeats (SSR) and single nucleotide polymorphisms (SNP) from both chloroplast genomes. Polycistronic mRNA was examined in L. acutangula and L. aegyptiaca using RNA sequences from Isoform sequencing to identify co-transcribed genes. IR size and locations were compared to other species and found to be relatively unchanged. Phylogenetic analysis confirmed the close relationship between L. acutangula and L. aegyptiaca in the Cucurbitaceae lineage and showed separation of the Luffa monophyletic clade from other species in the subtribe Sicyocae. The results obtained from this study can be useful for studying the evolution of Cucurbitaceae plants.

16.
PeerJ ; 8: e9608, 2020.
Article in English | MEDLINE | ID: mdl-33240580

ABSTRACT

Sugarcane contributes 80% of global sugar production and to bioethanol generation for the bioenergy industry. Its productivity is threatened by drought that can cause up to 60% yield loss. This study used RNA-Seq to gain a better understanding of the underlying mechanism by which drought-tolerant sugarcane copes with water stress. We compared gene expression in KPS01-12 (drought-tolerant genotype) and UT12 (drought-sensitive genotype) that have significantly different yield loss rates under drought conditions. We treated KPS01-12 and UT12 with mild and moderate water stress and found differentially expressed genes in various biological processes. KPS01-12 had higher expression of genes that were involved in water retention, antioxidant secondary metabolite biosynthesis, and oxidative and osmotic stress response than UT12. In contrast, the sensitive genotype had more down-regulated genes that were involved in photosynthesis, carbon fixation and Calvin cycle than the tolerant genotype. Our obtained expression profiles suggest that the tolerant sugarcane has a more effective genetic response than the sensitive genotype at the initiation of drought stress. The knowledge gained from this study may be applied in breeding programs to improve sugarcane production in drought conditions.

17.
PeerJ ; 8: e10340, 2020.
Article in English | MEDLINE | ID: mdl-33240651

ABSTRACT

Marine organisms are important to global food security as they are the largest source of animal proteins feeding mankind. Genomics-assisted aquaculture can increase yield while preserving the environment to ensure sufficient and sustainable production for global food security. However, only few high-quality genome sequences of marine organisms, especially shellfish, are available to the public partly because of the difficulty in the sequence assembly due to the complex nature of their genomes. A key step for a successful genome sequencing is the preparation of high-quality high molecular weight (HMW) genomic DNA. This study evaluated the effectiveness of five DNA extraction protocols (CTAB, Genomic-tip, Mollusc DNA, TIANamp Marine Animals DNA, and Sbeadex livestock kits) in obtaining shrimp HMW DNA for a long-read sequencing platform. DNA samples were assessed for quality and quantity using a Qubit fluorometer, NanoDrop spectrophotometer and pulsed-field gel electrophoresis. Among the five extraction methods examined without further optimization, the Genomic-tip kit yielded genomic DNA with the highest quality. However, further modifications of these established protocols might yield even better DNA quality and quantity. To further investigate whether the obtained genomic DNA could be used in a long-read sequencing application, DNA samples from the top three extraction methods (CTAB method, Genomic-tip and Mollusc DNA kits) were used for Pacific Biosciences (PacBio) library construction and sequencing. Genomic DNA obtained from Genomic-tip and Mollusc DNA kits allowed successful library construction, while the DNA obtained from the CTAB method did not. Genomic DNA isolated using the Genomic-tip kit yielded a higher number of long reads (N50 of 14.57 Kb) than those obtained from Mollusc DNA kits (N50 of 9.74 Kb). Thus, this study identified an effective extraction method for high-quality HMW genomic DNA of shrimp that can be applied to other marine organisms for a long-read sequencing platform.

18.
Sci Rep ; 10(1): 15980, 2020 10 07.
Article in English | MEDLINE | ID: mdl-33028920

ABSTRACT

We have assembled the complete sequence of the Durio zibethinus chloroplast genome using long PacBio reads. Durian is a valuable commercial tree that produces durian fruit, which is popular in Southeast Asia. The chloroplast genome assembled into a single 143 kb cyclic contig that contained 111 genes. There were 46 short direct repeats (45 to 586 bp) and five short inverted repeats (63 to 169 bp). The long reads that were used for the assembly span the entire chloroplast with > 10 kb overlaps and multiple long reads join the start of the contig to the end of the contig. The durian chloroplast was found to lack the large inverted repeat that is common in chloroplast genomes. An additional 24 durian varieties were sequenced and compared to the assembly and found to also lack the large inverted repeat. There were nine SNPs among the varieties.


Subject(s)
Bombacaceae/genetics , Chloroplasts/genetics , Evolution, Molecular , Fruit/genetics , Genome, Chloroplast , Phylogeny , Sequence Analysis, DNA/methods , Bombacaceae/classification , Bombacaceae/growth & development , Fruit/growth & development
19.
Plants (Basel) ; 9(9)2020 Sep 21.
Article in English | MEDLINE | ID: mdl-32967378

ABSTRACT

Vigna mungo is cultivated in approximately 5 million hectares worldwide. The chloroplast genome of this species has not been previously reported. In this study, we sequenced the genome and transcriptome of the V. mungo chloroplast. We identified many positively selected genes in the photosynthetic pathway (e.g., rbcL, ndhF, and atpF) and RNA polymerase genes (e.g., rpoC2) from the comparison of the chloroplast genome of V. mungo, temperate legume species, and tropical legume species. Our transcriptome data from PacBio isoform sequencing showed that the 51-kb DNA inversion could affect the transcriptional regulation of accD polycistronic. Using Illumina deep RNA sequencing, we found RNA editing of clpP in the leaf, shoot, flower, fruit, and root tissues of V. mungo. We also found three G-to-A RNA editing events that change guanine to adenine in the transcripts transcribed from the adenine-rich regions of the ycf4 gene. The edited guanine bases were found particularly in the chloroplast genome of the Vigna species. These G-to-A RNA editing events were likely to provide a mechanism for correcting DNA base mutations. The V. mungo chloroplast genome sequence and the analysis results obtained in this study can apply to phylogenetic studies and chloroplast genome engineering.

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