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Culture-independent-based analyses identified site-specific microbial in two oil-wells from Malaysia

Cheng-Yee Fish-Low, Chee HY, Ainon Hamzah

Sains Malaysiana, 2015;44:1625-1633.

Microbial communities of two oil reservoirs from Malaysia, denoted as Platform Bo and Platform Pe were studied using
culture-independent approach. Environmental DNA was extracted and the universal amplified ribosomal region (UARR)
was target amplified for both prokaryotes and eukaryotes. The amplified products were purified and cloned into pTZ57R/T
vector to construct the 16S/18S rDNA library. Restriction endocucleases HhaI and MspI were used to screen the library.
From that, 125 and 253 recombinant plasmid representative clones from Platform Bo and Platform Pe, respectively, were
sent for DNA sequencing. Twenty-six operational taxonomic units (OTUs) consist of 20 genera detected at Platform Bo
and 17 OTUs consist of 13 genera detected at Platform Pe. Marinobacter and Acinetobacter species co-occurred in both
platforms whereas the rest are site-specific. Gammaproteobacteria accounted for 86.0% of the microbial community in
Platform Bo, where OTUs affiliated to Marinobacter, Pseudomonas and Marinobacterium that were the most abundant. The
major OTUs in the Platform Pe were with affinities to Achromobacter, followed by Stenotrophomonas and Serratia. The
only archaeal isolates were detected in Platform Pe, which affiliated to Thermocladium. The singletons and doubletons
accounted for about 50.0% of the OTU abundance in both platforms, which considerably significant despite their rare
occurrence.

Matched MeSH terms: Gene Library
Fulltext DNA-based taxonomy of a mangrove-associated community of fishes in Southeast Asia

Zainal Abidin DH, Mohd Nor SA, Lavoué S, A Rahim M, Jamaludin NA, Mohammed Akib NA

Sci Rep, 2021 Sep 07;11(1):17800.
PMID: 34493747 DOI: 10.1038/s41598-021-97324-1

The Merbok Estuary comprises one of the largest remaining mangrove forests in Peninsular Malaysia. Its value is significant as it provides important services to local and global communities. It also offers a unique opportunity to study the structure and functioning of mangrove ecosystems. However, its biodiversity is still partially inventoried, limiting its research value. A recent checklist based on morphological examination, reported 138 fish species residing, frequenting or subject to entering the Merbok Estuary. In this work, we reassessed the fish diversity of the Merbok Estuary by DNA barcoding 350 specimens assignable to 134 species initially identified based on morphology. Our results consistently revealed the presence of 139 Molecular Operational Taxonomic Units (MOTUs). 123 of them are congruent with morphology-based species delimitation (one species = one MOTU). In two cases, two morphological species share the same MOTU (two species = one MOTU), while we unveiled cryptic diversity (i.e. COI-based genetic variability > 2%) within seven other species (one species = two MOTUs), calling for further taxonomic investigations. This study provides a comprehensive core-list of fish taxa in Merbok Estuary, demonstrating the advantages of combining morphological and molecular evidence to describe diverse but still poorly studied tropical fish communities. It also delivers a large DNA reference collection for brackish fishes occurring in this region which will facilitate further biodiversity-oriented research studies and management activities.

Matched MeSH terms: Gene Library
Fulltext Towards complete and error-free genome assemblies of all vertebrate species

Rhie A, McCarthy SA, Fedrigo O, Damas J, Formenti G, Koren S, et al.

Nature, 2021 Apr;592(7856):737-746.
PMID: 33911273 DOI: 10.1038/s41586-021-03451-0

High-quality and complete reference genome assemblies are fundamental for the application of genomics to biology, disease, and biodiversity conservation. However, such assemblies are available for only a few non-microbial species1-4. To address this issue, the international Genome 10K (G10K) consortium5,6 has worked over a five-year period to evaluate and develop cost-effective methods for assembling highly accurate and nearly complete reference genomes. Here we present lessons learned from generating assemblies for 16 species that represent six major vertebrate lineages. We confirm that long-read sequencing technologies are essential for maximizing genome quality, and that unresolved complex repeats and haplotype heterozygosity are major sources of assembly error when not handled correctly. Our assemblies correct substantial errors, add missing sequence in some of the best historical reference genomes, and reveal biological discoveries. These include the identification of many false gene duplications, increases in gene sizes, chromosome rearrangements that are specific to lineages, a repeated independent chromosome breakpoint in bat genomes, and a canonical GC-rich pattern in protein-coding genes and their regulatory regions. Adopting these lessons, we have embarked on the Vertebrate Genomes Project (VGP), an international effort to generate high-quality, complete reference genomes for all of the roughly 70,000 extant vertebrate species and to help to enable a new era of discovery across the life sciences.

Matched MeSH terms: Gene Library
Fulltext RNA-seq and ChIP-seq as Complementary Approaches for Comprehension of Plant Transcriptional Regulatory Mechanism

Muhammad II, Kong SL, Akmar Abdullah SN, Munusamy U

Int J Mol Sci, 2019 Dec 25;21(1).
PMID: 31881735 DOI: 10.3390/ijms21010167

The availability of data produced from various sequencing platforms offer the possibility to answer complex questions in plant research. However, drawbacks can arise when there are gaps in the information generated, and complementary platforms are essential to obtain more comprehensive data sets relating to specific biological process, such as responses to environmental perturbations in plant systems. The investigation of transcriptional regulation raises different challenges, particularly in associating differentially expressed transcription factors with their downstream responsive genes. In this paper, we discuss the integration of transcriptional factor studies through RNA sequencing (RNA-seq) and Chromatin Immunoprecipitation sequencing (ChIP-seq). We show how the data from ChIP-seq can strengthen information generated from RNA-seq in elucidating gene regulatory mechanisms. In particular, we discuss how integration of ChIP-seq and RNA-seq data can help to unravel transcriptional regulatory networks. This review discusses recent advances in methods for studying transcriptional regulation using these two methods. It also provides guidelines for making choices in selecting specific protocols in RNA-seq pipelines for genome-wide analysis to achieve more detailed characterization of specific transcription regulatory pathways via ChIP-seq.

Matched MeSH terms: Gene Library
Fulltext Determination of genetic aberrations and novel transcripts involved in the pathogenesis of oligodendroglioma using array comparative genomic hybridization and next generation sequencing

Hassanudin SA, Ponnampalam SN, Amini MN

Oncol Lett, 2019 Feb;17(2):1675-1687.
PMID: 30675227 DOI: 10.3892/ol.2018.9811

The aim of the present study was to determine the genetic aberrations and novel transcripts, particularly the fusion transcripts, involved in the pathogenesis of low-grade and anaplastic oligodendroglioma. In the present study, tissue samples were obtained from patients with oligodendroglioma and additionally from archived tissue samples from the Brain Tumor Tissue Bank of the Brain Tumor Foundation of Canada. Six samples were obtained, three of which were low-grade oligodendroglioma and the other three anaplastic oligodendroglioma. DNA and RNA were extracted from each tissue sample. The resulting genomic DNA was then hybridized using the Agilent CytoSure 4×180K oligonucleotide array. Human reference DNA and samples were labeled using Cy3 cytidine 5'-triphosphate (CTP) and Cy5 CTP, respectively, while human Cot-1 DNA was used to reduce non-specific binding. Microarray-based comparative genomic hybridization data was then analyzed for genetic aberrations using the Agilent Cytosure Interpret software v3.4.2. The total RNA isolated from each sample was mixed with oligo dT magnetic beads to enrich for poly(A) mRNA. cDNAs were then synthesized and subjected to end-repair, poly(A) addition and connected using sequencing adapters using the Illumina TruSeq RNA Sample Preparation kit. The fragments were then purified and selected as templates for polymerase chain reaction amplification. The final library was constructed with fragments between 350-450 base pairs and sequenced using deep transcriptome sequencing on an Illumina HiSeq 2500 sequencer. The array comparative genomic hybridization revealed numerous amplifications and deletions on several chromosomes in all samples. However, the most interesting result was from the next generation sequencing, where one anaplastic oligodendroglioma sample was demonstrated to have five novel fusion genes that may potentially serve a critical role in tumor pathogenesis and progression.

Matched MeSH terms: Gene Library
Fulltext Analysis of the leaf transcriptome of Musa acuminata during interaction with Mycosphaerella musicola: gene assembly, annotation and marker development

Passos MA, de Cruz VO, Emediato FL, de Teixeira CC, Azevedo VC, Brasileiro AC, et al.

BMC Genomics, 2013 Feb 05;14:78.
PMID: 23379821 DOI: 10.1186/1471-2164-14-78

BACKGROUND: Although banana (Musa sp.) is an important edible crop, contributing towards poverty alleviation and food security, limited transcriptome datasets are available for use in accelerated molecular-based breeding in this genus. 454 GS-FLX Titanium technology was employed to determine the sequence of gene transcripts in genotypes of Musa acuminata ssp. burmannicoides Calcutta 4 and M. acuminata subgroup Cavendish cv. Grande Naine, contrasting in resistance to the fungal pathogen Mycosphaerella musicola, causal organism of Sigatoka leaf spot disease. To enrich for transcripts under biotic stress responses, full length-enriched cDNA libraries were prepared from whole plant leaf materials, both uninfected and artificially challenged with pathogen conidiospores.
RESULTS: The study generated 846,762 high quality sequence reads, with an average length of 334 bp and totalling 283 Mbp. De novo assembly generated 36,384 and 35,269 unigene sequences for M. acuminata Calcutta 4 and Cavendish Grande Naine, respectively. A total of 64.4% of the unigenes were annotated through Basic Local Alignment Search Tool (BLAST) similarity analyses against public databases.Assembled sequences were functionally mapped to Gene Ontology (GO) terms, with unigene functions covering a diverse range of molecular functions, biological processes and cellular components. Genes from a number of defense-related pathways were observed in transcripts from each cDNA library. Over 99% of contig unigenes mapped to exon regions in the reference M. acuminata DH Pahang whole genome sequence. A total of 4068 genic-SSR loci were identified in Calcutta 4 and 4095 in Cavendish Grande Naine. A subset of 95 potential defense-related gene-derived simple sequence repeat (SSR) loci were validated for specific amplification and polymorphism across M. acuminata accessions. Fourteen loci were polymorphic, with alleles per polymorphic locus ranging from 3 to 8 and polymorphism information content ranging from 0.34 to 0.82.
CONCLUSIONS: A large set of unigenes were characterized in this study for both M. acuminata Calcutta 4 and Cavendish Grande Naine, increasing the number of public domain Musa ESTs. This transcriptome is an invaluable resource for furthering our understanding of biological processes elicited during biotic stresses in Musa. Gene-based markers will facilitate molecular breeding strategies, forming the basis of genetic linkage mapping and analysis of quantitative trait loci.

Matched MeSH terms: Gene Library
Diversity of bovine rumen methanogens In vitro in the presence of condensed tannins, as determined by sequence analysis of 16S rRNA gene library

Tan HY, Sieo CC, Lee CM, Abdullah N, Liang JB, Ho YW

J Microbiol, 2011 Jun;49(3):492-8.
PMID: 21717338 DOI: 10.1007/s12275-011-0319-7

Molecular diversity of rumen archaeal populations from bovine rumen fluid incubated with or without condensed tannins was investigated using 16S rRNA gene libraries. The predominant order of rumen archaea in the 16S rRNA gene libraries of the control and condensed tannins treatment was found to belong to a novel group of rumen archaea that is distantly related to the order Thermoplasmatales, with 59.5% (15 phylotypes) and 81.43% (21 phylotypes) of the total clones from the control and treatment clone libraries, respectively. The 16S rRNA gene library of the control was found to have higher proportions of methanogens from the orders Methanomicrobiales (32%) and Methanobacteriales (8.5%) as compared to those found in the condensed tannins treatment clone library in both orders (16.88% and 1.68% respectively). The phylotype distributed in the order Methanosarcinales was only found in the control clone library. The study indicated that condensed tannins could alter the diversity of bovine rumen methanogens.

Matched MeSH terms: Gene Library*
Production of 3-Oxo-2-(2'-pentenyl)-cyclopentane-1-octanoic Acid in the Fungus Aspergillus oryzae: A Step Towards Heterologous Production of Pyrethrins in Fungi

Mohamed ME, Pahirulzaman KA, Lazarus CM

Mol Biotechnol, 2016 Mar;58(3):172-8.
PMID: 26718544 DOI: 10.1007/s12033-015-9911-0

Pyrethrins are natural insecticides, which accumulate to high concentrations in pyrethrum (Chrysanthemum cinerariaefolium) flowers. Synthetic pyrethroids are more stable, more efficacious and cheaper, but contemporary requirements for safe and environmentally friendly pesticides encourage a return to the use of natural pyrethrins, and this would be favoured by development of an efficient route to their production by microbial fermentation. The biosynthesis of pyrethrins involves ester linkage between an acid moiety (chrysanthemoyl or pyrethroyl, synthesised via the mevalonic acid pathway from glucose), and an alcohol (pyrethrolone). Pyrethrolone is generated from 3-oxo-2-(2'-pentenyl)-cyclopentane-1-octanoic acid, which originates from α-linolenic acid via the jasmonic acid biosynthetic cascade. The first four genes in this cascade, encoding lipoxygenase 2, allene-oxide synthase, allene-oxide cyclase 2 and 12-oxophytodienoic acid reductase 3, were amplified from an Arabidopsis thaliana cDNA library, cloned in a purpose-built fungal multigene expression vector and expressed in Aspergillus oryzae. HPLC-MS analysis of the transgenic fungus homogenate gave good evidence for the presence of 3-oxo-2-(2'-pentenyl)-cyclopentane-1-octanoic acid.

Matched MeSH terms: Gene Library
Fulltext Protein-protein interactions between A. aegypti midgut and dengue virus 2: two-hybrid screens using the midgut cDNA library

Tham HW, Balasubramaniam VR, Chew MF, Ahmad H, Hassan SS

J Infect Dev Ctries, 2015 Dec 30;9(12):1338-49.
PMID: 26719940 DOI: 10.3855/jidc.6422

INTRODUCTION: Dengue virus (DENV) is principally transmitted by the Aedes aegypti mosquito. To date, mosquito population control remains the key strategy for reducing the continuing spread of DENV. The focus on the development of new vector control strategies through an understanding of the mosquito-virus relationship is essential, especially targeting the midgut, which is the first mosquito organ exposed to DENV infection.
METHODOLOGY: A cDNA library derived from female adult A. aegypti mosquito midgut cells was established using the switching mechanism at the 5' end of the RNA transcript (SMART), in combination with a highly potent recombination machinery of Saccharomyces cerevisiae. Gal4-based yeast two-hybrid (Y2H) assays were performed against DENV-2 proteins (E, prM, M, and NS1). Mammalian two-hybrid (M2H) and double immunofluorescence assays (IFA) were conducted to validate the authenticity of the three selected interactions.
RESULTS: The cDNA library was of good quality based on its transformation efficiency, cell density, titer, and the percentage of insert size. A total of 36 midgut proteins interacting with DENV-2 proteins were identified, some involved in nucleic acid transcription, oxidoreductase activity, peptidase activity, and ion binding. Positive outcomes were obtained from the three selected interactions validated using M2H and double IFA assays.
CONCLUSIONS: The identified proteins have different biological activities that may aid in the virus replication pathway. Therefore, the midgut cDNA library is a valuable tool for identifying DENV-2 interacting proteins. The positive outcomes of the three selected proteins validated supported the quality of the cDNA library and the robustness of the Y2H mechanisms.

Matched MeSH terms: Gene Library
Fulltext Potential immunogenic polypeptides of Burkholderia pseudomallei identified by shotgun expression library and evaluation of their efficacy for serodiagnosis of melioidosis

Puah SM, Puthucheary SD, Chua KH

Int J Med Sci, 2013;10(5):539-47.
PMID: 23532805 DOI: 10.7150/ijms.5516

The search for novel immunogenic polypeptides to improve the accuracy and reliability of serologic diagnostic methods for Burkholderia pseudomallei infection is ongoing. We employed a rapid and efficient approach to identify such polypeptides with sera from melioidosis patients using a small insert genomic expression library created from clinically confirmed local virulent isolates of B. pseudomallei. After 2 rounds of immunoscreening, 6 sero-positive clones expressing immunogenic peptides were sequenced and their identities were: benzoate 1,2-dioxygenase beta subunit, a putative 200 kDa antigen p200, phosphotransferase enzyme family protein, short chain dehydrogenase and 2 hypothetical proteins. These immunogens were then transferred to an ELISA platform for further large scale screening. By combining shotgun expression library and ELISA assays, we identified 2 polypeptides BPSS1904 (benzoate 1,2-dioxygenase beta subunit) and BPSL3130 (hypothetical protein), which had sensitivities of 78.9% and 79.4% and specificities of 88.1% and 94.8%, respectively in ELISA test, thus suggesting that both are potential candidate antigens for the serodiagnosis of infections caused by B. pseudomallei.

Matched MeSH terms: Gene Library
Identification of Toxoplasma gondii in-vivo induced antigens by cDNA library immunoscreening with chronic toxoplasmosis sera

Amerizadeh A, Idris ZM, Khoo BY, Kotresha D, Yunus MH, Karim IZ, et al.

Microb Pathog, 2013 Jan;54:60-6.
PMID: 23044055 DOI: 10.1016/j.micpath.2012.09.006

Toxoplasmosis is an infection caused by the parasite Toxoplasma gondii. Chronically-infected individuals with a compromised immune system are at risk for reactivation of the disease. In-vivo induced antigen technology (IVIAT) is a promising method for the identification of antigens expressed in-vivo. The aim of the present study was to apply IVIAT to identify antigens which are expressed in-vivo during T. gondii infection using sera from individuals with chronic toxoplasmosis. Forty serum samples were pooled, pre-adsorped against three different preparations of antigens, from each in-vitro grown T. gondii and Escherichia coli XLBlue MRF', and then used to screen a T. gondii cDNA expression library. Sequencing of DNA inserts from positive clones showed eight open reading frames with high homology to T. gondii genes. Expression analysis using quantitative real-time PCR showed that SAG1-related sequence 3 (SRS3) and two hypothetical genes were up-regulated in-vivo relative to their expression levels in-vitro. These three proteins also showed high sensitivity and specificity when tested with individual serum samples. Five other proteins namely M16 domain peptidase, microneme protein, elongation factor 1-alpha, pre-mRNA-splicing factor and small nuclear ribonucleoprotein F had lower RNA expression in-vivo as compared to in-vitro. SRS3 and the two hypothetical proteins warrant further investigation into their roles in the pathogenesis of toxoplasmosis.

Matched MeSH terms: Gene Library
Fulltext Transcriptome analysis of the Cryptocaryon irritans tomont stage identifies potential genes for the detection and control of cryptocaryonosis

Lokanathan Y, Mohd-Adnan A, Wan KL, Nathan S

BMC Genomics, 2010;11:76.
PMID: 20113487 DOI: 10.1186/1471-2164-11-76

Cryptocaryon irritans is a parasitic ciliate that causes cryptocaryonosis (white spot disease) in marine fish. Diagnosis of cryptocaryonosis often depends on the appearance of white spots on the surface of the fish, which are usually visible only during later stages of the disease. Identifying suitable biomarkers of this parasite would aid the development of diagnostic tools and control strategies for C. irritans. The C. irritans genome is virtually unexplored; therefore, we generated and analyzed expressed sequence tags (ESTs) of the parasite to identify genes that encode for surface proteins, excretory/secretory proteins and repeat-containing proteins.

Matched MeSH terms: Gene Library
Discovery of precursor and mature microRNAs and their putative gene targets using high-throughput sequencing in pineapple (Ananas comosus var. comosus)

Yusuf NH, Ong WD, Redwan RM, Latip MA, Kumar SV

Gene, 2015 Oct 15;571(1):71-80.
PMID: 26115767 DOI: 10.1016/j.gene.2015.06.050

MicroRNAs (miRNAs) are a class of small, endogenous non-coding RNAs that negatively regulate gene expression, resulting in the silencing of target mRNA transcripts through mRNA cleavage or translational inhibition. MiRNAs play significant roles in various biological and physiological processes in plants. However, the miRNA-mediated gene regulatory network in pineapple, the model tropical non-climacteric fruit, remains largely unexplored. Here, we report a complete list of pineapple mature miRNAs obtained from high-throughput small RNA sequencing and precursor miRNAs (pre-miRNAs) obtained from ESTs. Two small RNA libraries were constructed from pineapple fruits and leaves, respectively, using Illumina's Solexa technology. Sequence similarity analysis using miRBase revealed 579,179 reads homologous to 153 miRNAs from 41 miRNA families. In addition, a pineapple fruit transcriptome library consisting of approximately 30,000 EST contigs constructed using Solexa sequencing was used for the discovery of pre-miRNAs. In all, four pre-miRNAs were identified (MIR156, MIR399, MIR444 and MIR2673). Furthermore, the same pineapple transcriptome was used to dissect the function of the miRNAs in pineapple by predicting their putative targets in conjunction with their regulatory networks. In total, 23 metabolic pathways were found to be regulated by miRNAs in pineapple. The use of high-throughput sequencing in pineapples to unveil the presence of miRNAs and their regulatory pathways provides insight into the repertoire of miRNA regulation used exclusively in this non-climacteric model plant.

Matched MeSH terms: Gene Library
Identification of chromosome 21 materials using the whole chromosome 21 specific library

Isa MN, Boyd E, Turner TL, Tolmie J, Connor JM

Southeast Asian J. Trop. Med. Public Health, 1995;26 Suppl 1:92-5.
PMID: 8629150

The chromosome in situ suppression hybridization or chromosome painting technic was applied to confirm and eliminate the markers involving chromosome 21 segments using a chromosome 21 DNA library. The library ATCCLL21SNO2 was amplified, directly biotinylated using the polymerase chain reaction. The results demonstrated a translocation of chromosome 21 material on chromosome 2 and X and eliminate the origin of the marker. Thus, the technique provides an important tool to complement the conventional G-banding technic.

Matched MeSH terms: Gene Library
Fulltext Haplotype divergence and multiple candidate genes at Rphq2, a partial resistance QTL of barley to Puccinia hordei

Yeo FK, Wang Y, Vozabova T, Huneau C, Leroy P, Chalhoub B, et al.

Theor Appl Genet, 2016 Feb;129(2):289-304.
PMID: 26542283 DOI: 10.1007/s00122-015-2627-5

Rphq2, a minor gene for partial resistance to Puccinia hordei , was physically mapped in a 188 kbp introgression with suppressed recombination between haplotypes of rphq2 and Rphq2 barley cultivars.

Matched MeSH terms: Gene Library
Structural and functional changes with depth in microbial communities in a tropical Malaysian peat swamp forest

Jackson CR, Liew KC, Yule CM

Microb Ecol, 2009 Apr;57(3):402-12.
PMID: 18548182 DOI: 10.1007/s00248-008-9409-4

Tropical peat swamp forests are important and endangered ecosystems, although little is known of their microbial diversity and ecology. We used molecular and enzymatic techniques to examine patterns in prokaryotic community structure and overall microbial activity at 0-, 10-, 20-, and 50-cm depths in sediments in a peat swamp forest in Malaysia. Denaturing gradient gel electrophoresis profiles of amplified 16S ribosomal ribonucleic acid (rRNA) gene fragments showed that different depths harbored different bacterial assemblages and that Archaea appeared to be limited to the deeper samples. Cloning and sequencing of longer 16S rRNA gene fragments suggested reduced microbial diversity in the deeper samples compared to the surface. Bacterial clone libraries were largely dominated by ribotypes affiliated with the Acidobacteria, which accounted for at least 27-54% of the sequences obtained. All of the sequenced representatives from the archaeal clone libraries were Crenarchaeota. Activities of microbial extracellular enzymes involved in carbon, nitrogen, and phosphorus cycling declined appreciably with depth, the only exception being peroxidase. These results show that tropical peat swamp forests are unusual systems with microbial assemblages dominated by members of the Acidobacteria and Crenarchaeota. Microbial communities show clear changes with depth, and most microbial activity is likely confined to populations in the upper few centimeters, the site of new leaf litter fall, rather than the deeper, older, peat layers.

Matched MeSH terms: Gene Library
Regional and accelerated molecular evolution in group I snake venom gland phospholipase A2 isozymes

Chuman Y, Nobuhisa I, Ogawa T, Deshimaru M, Chijiwa T, Tan NH, et al.

Toxicon, 2000 Mar;38(3):449-62.
PMID: 10669032

In accordance with detection of a few phospholipase A2 (PLA2) isozyme genes by Southern blot analysis, only two cDNAs, named NnkPLA-I , and NnkPLA-II, encoding group I PLA2s, NnkPLA-I and NnkPLA-II, respectively, were isolated from the venom gland cDNA library of Elapinae Naja naja kaouthia of Malaysia. NnkPLA-I and NnkPLA-II showed four amino acid substitutions, all of which were brought about by single nucleotide substitution. No existence of clones encoding CM-II and CM-III, PLA2 isozymes which had been isolated from the venom of N. naja kaouthia of Thailand, in Malaysian N. naja kaouthia venom gland cDNA library was verified by dot blot hybridization analysis with particular probes. NnkPLA-I and NnkPLA-II differed from CM-II and CM-III with four and two amino acid substitutions, respectively, suggesting that their molecular evolution is regional. The comparison of NnkPLA-I, NnkPLA-II and cDNAs encoding other group I snake venom gland PLA2s indicated that the 5'- and 3'-untranslated regions are more conserved than the mature protein-coding region and that the number of nucleotide substitutions per nonsynonymous site is almost equal to that per synonymous site in the protein-coding region, suggesting that accelerated evolution has occurred in group I venom gland PLA2s possibly to acquire new physiological functions.

Matched MeSH terms: Gene Library
Fulltext Differential expression patterns of leukaemia associated genes in leukaemia cell lines compared to healthy controls

Ang Pei-Shen, Rajesh Ramasamy, Noor Hamidah Hussin, Cheong Soon-Keng, Seow Heng-Fong, Maha Abdullah

Malaysian Journal of Medicine and Health Sciences, 2016;12(1):0-0.
MyJurnal

Introduction: The phenotype and genotype of cancer cells portray hallmarks of cancer which may
have clinical value. Cancer cell lines are ideal models to study and confirm these characteristics. We
previously established two subtracted cDNA libraries with differentially expressed genes from an
acute myeloid leukaemia patient with poor prognosis (PP) and good prognosis (GP). Objective: To
compare gene expression of the leukaemia associated genes with selected biological characteristics
in leukaemia cell lines and normal controls. Methodology: Expression of 28 PP genes associated
with early fetal/embryonic development, HOX-related genes, hematopoiesis and aerobic glycolysis/
hypoxia genes and 36 GP genes involved in oxidative phosphorylation, protein synthesis, chromatin
remodelling and cell motility were examined in B-lymphoid (BV173, Reh and RS4;11) and myeloid
(HL-60, K562) leukaemia cell lines after 72h in culture as well as peripheral blood mononuclear cells
from healthy controls (N=5) using semi-quantitative polymerase chain reaction (PCR) method. Cell
cycle profiles were analysed on flow cytometry while MTT cytotoxicity assay was used to determine
drug resistance to epirubicin. Results: Genes expressed significantly higher in B-lymphoid leukaemia
cell lines compared to healthy controls were mostly of the GP library i.e. oxidative phosphorylation
(3/10), protein synthesis (4/11), chromatin remodelling (3/3) and actin cytoskeleton genes (1/5). Only
two genes with significant difference were from the PP library. Cancer associated genes, HSPA9 and
PSPH (GP library) and BCAP31 (PP library) were significantly higher in the B-lymphoid leukemia cell
lines. No significant difference was observed between myeloid cell lines and healthy controls. This
may also be due heterogeneity of cell lines studied. PBMC from healthy controls were not in cell cycle.
G2/M profiles and growth curves showed B-lymphoid cells just reaching plateau after 72 hour culture
while myeloid cells were declining. IC50 values from cytotoxicity assay revealed myeloid cell lines had
an average 13-fold higher drug resistance to epirubicin compared to B-lymphoid cell lines. Only CCL1,
was expressed at least two-fold higher in myeloid compared to B-lymphoid cell lines. In contrast,
MTRNR2, EEF1A1, PTMA, HLA-DR, C6orf115, PBX3, ENPP4, SELL, and IL3Ra were expressed
more than 2-fold higher in B-lymphoid compared to myeloid cell lines studied here. Conclusion: Thus,
B-lymphoid leukaemia cell lines here exhibited active, proliferating characteristics closer to GP genes.
Higher expression of several genes in B-lymphoid compared to myeloid leukaemia cell lines may be
useful markers to study biological differences including drug resistance between lineages.

Matched MeSH terms: Gene Library
Fulltext Oil palm (Elaeis guineensis Jacq.) tissue culture ESTs: identifying genes associated with callogenesis and embryogenesis

Low ET, Alias H, Boon SH, Shariff EM, Tan CY, Ooi LC, et al.

BMC Plant Biol, 2008 May 29;8:62.
PMID: 18507865 DOI: 10.1186/1471-2229-8-62

BACKGROUND: Oil palm (Elaeis guineensis Jacq.) is one of the most important oil bearing crops in the world. However, genetic improvement of oil palm through conventional breeding is extremely slow and costly, as the breeding cycle can take up to 10 years. This has brought about interest in vegetative propagation of oil palm. Since the introduction of oil palm tissue culture in the 1970s, clonal propagation has proven to be useful, not only in producing uniform planting materials, but also in the development of the genetic engineering programme. Despite considerable progress in improving the tissue culture techniques, the callusing and embryogenesis rates from proliferating callus cultures remain very low. Thus, understanding the gene diversity and expression profiles in oil palm tissue culture is critical in increasing the efficiency of these processes.
RESULTS: A total of 12 standard cDNA libraries, representing three main developmental stages in oil palm tissue culture, were generated in this study. Random sequencing of clones from these cDNA libraries generated 17,599 expressed sequence tags (ESTs). The ESTs were analysed, annotated and assembled to generate 9,584 putative unigenes distributed in 3,268 consensi and 6,316 singletons. These unigenes were assigned putative functions based on similarity and gene ontology annotations. Cluster analysis, which surveyed the relatedness of each library based on the abundance of ESTs in each consensus, revealed that lipid transfer proteins were highly expressed in embryogenic tissues. A glutathione S-transferase was found to be highly expressed in non-embryogenic callus. Further analysis of the unigenes identified 648 non-redundant simple sequence repeats and 211 putative full-length open reading frames.
CONCLUSION: This study has provided an overview of genes expressed during oil palm tissue culture. Candidate genes with expression that are modulated during tissue culture were identified. However, in order to confirm whether these genes are suitable as early markers for embryogenesis, the genes need to be tested on earlier stages of tissue culture and a wider range of genotypes. This collection of ESTs is an important resource for genetic and genome analyses of the oil palm, particularly during tissue culture development.

Matched MeSH terms: Gene Library
Effect of condensed tannins on bovine rumen protist diversity based on 18S rRNA gene sequences

Tan HY, Sieo CC, Abdullah N, Liang JB, Huang XD, Ho YW

J. Eukaryot. Microbiol., 2013 Jan-Feb;60(1):98-100.
PMID: 23205499 DOI: 10.1111/jeu.12011

Molecular diversity of protists from bovine rumen fluid incubated with condensed tannins of Leucaena leucocephala hybrid-Rendang at 20 mg/500 mg dry matter (treatment) or without condensed tannins (control) was investigated using 18S rRNA gene library. Clones from the control library were distributed within nine genera, but clones from the condensed tannin treatment clone library were related to only six genera. Diversity estimators such as abundance-based coverage estimation and Chao1 showed significant differences between the two libraries, although no differences were found based on Shannon-Weaver index and Libshuff.

Matched MeSH terms: Gene Library

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