Displaying publications 1 - 20 of 65 in total

  1. Omar N, Lim TS
    Methods Mol Biol, 2018;1701:25-44.
    PMID: 29116498 DOI: 10.1007/978-1-4939-7447-4_2
    This protocol describes the processes involved in the generation of human antibody libraries in Fab format. The antibody repertoire is derived from peripheral blood mononucleocytes focusing on different immunoglobulin isotypes. A two-step cloning process was used to generate a diverse human Fab library for subsequent selection by phage display. The method can be applied for the generation of both naive and immune antibody libraries. The naive repertoire allows for the library to be applied for the generation of human monoclonal antibodies against a broad range of target antigens making it a useful resource for antibody generation. However, the immune repertoire will be focused against target antigens from a particular disease. The protocol will focus on the generation of the library including the panning process.
    Matched MeSH terms: Gene Library*
  2. Hairul Bahara NH, Tye GJ, Choong YS, Ong EB, Ismail A, Lim TS
    Biologicals, 2013 Jul;41(4):209-16.
    PMID: 23647952 DOI: 10.1016/j.biologicals.2013.04.001
    With major developments in molecular biology, numerous display technologies have been successfully introduced for recombinant antibody production. Even so, phage display still remains the gold standard for recombinant antibody production. Its success is mainly attributed to the robust nature of phage particles allowing for automation and adaptation to modifications. The generation of monospecific binders provides a vital tool for diagnostics at a lower cost and higher efficiency. The flexibility to modify recombinant antibodies allows great applicability to various platforms for use. This review presents phage display technology, application and modifications of recombinant antibodies for diagnostics.
    Matched MeSH terms: Gene Library*
  3. Usman AR, Khandaker MU, Haba H, Otuka N, Murakami M, Komori Y
    Appl Radiat Isot, 2016 08;114:104-13.
    PMID: 27227905 DOI: 10.1016/j.apradiso.2016.04.027
    The excitation functions were measured for the (nat)Cu(α,x)(66,67)Ga,(65)Zn,(57,58,60)Co reactions in the energy range of 16.5 -50MeV. A conventional stacked-foil activation technique combined with HPGe γ-ray spectrometry was employed to determine cross-sections. The measured cross-sections were critically compared with relevant previous experimental data and also with the evaluated data in the TENDL-2014 library. Present results confirmed some of the previous experimental data, whereas only a partial agreement was found with the evaluated data. The measured data are useful for reducing the existing discrepancies in the literature, to improve the nuclear reaction model codes, and to enrich the experimental database towards various applications.
    Matched MeSH terms: Gene Library
  4. Thanh T, Chi VT, Abdullah MP, Omar H, Noroozi M, Ky H, et al.
    Mol Biol Rep, 2011 Jan;38(1):177-82.
    PMID: 20354903 DOI: 10.1007/s11033-010-0092-4
    Green microalga Ankistrodesmus convolutus Corda is a fast growing alga which produces appreciable amount of carotenoids and polyunsaturated fatty acids. To our knowledge, this is the first report on the construction of cDNA library and preliminary analysis of ESTs for this species. The titers of the primary and amplified cDNA libraries were 1.1×10(6) and 6.0×10(9) pfu/ml respectively. The percentage of recombinants was 97% in the primary library and a total of 337 out of 415 original cDNA clones selected randomly contained inserts ranging from 600 to 1,500 bps. A total of 201 individual ESTs with sizes ranging from 390 to 1,038 bps were then analyzed and the BLASTX score revealed that 35.8% of the sequences were classified as strong match, 38.3% as nominal and 25.9% as weak match. Among the ESTs with known putative function, 21.4% of them were found to be related to gene expression, 14.4% ESTs to photosynthesis, 10.9% ESTs to metabolism, 5.5% ESTs to miscellaneous, 2.0% to stress response, and the remaining 45.8% were classified as novel genes. Analysis of ESTs described in this paper can be an effective approach to isolate and characterize new genes from A. convolutus and thus the sequences obtained represented a significant contribution to the extensive database of sequences from green microalgae.
    Matched MeSH terms: Gene Library*
  5. Shah FH, Cha TS
    Plant Sci, 2000 May 29;154(2):153-160.
    PMID: 10729614
    The differential display method was used to isolate cDNAs corresponding to transcripts that accumulate during the period of lipid synthesis, 12-20 weeks after anthesis (WAA) in the mesocarp of two oil palms, Elaeis oleifera and Elaeis guineensis, Tenera. DNA-free total RNA from mesocarp and kernel of E. guineensis, Tenera and E. oleifera (15 WAA) were used to obtain differential gene expression patterns between these tissues from the two species. In this report, we describe the isolation and characterization of a specific cDNA clone, MO1 (434 bp) which was shown to be mesocarp-specific as well as species-specific for E. oleifera Sequencing of this fragment showed homology to the enzyme sesquiterpene synthase. Its longer cDNA clone, pMO1 (1072 bp), isolated from a 15-week E. oleifera mesocarp cDNA library confirmed that it encodes for sesquiterpene synthase. The complete sequence of 1976 bp was obtained using 5'RACE method. Northern hybridization showed that MO1 and pMO1 mRNA transcripts are highly expressed only in the mesocarp of E. oleifera from 5 to 20 WAA. No expression was detected in the kernel (12-17 WAA) and vegetative tissues of both species nor in the mesocarp of E. guineensis. This is the first communication to document on the isolation and characterisation of a mesocarp-and species-specific cDNA clone from oil palm.
    Matched MeSH terms: Gene Library
  6. Farah Fadwa Benbelgacem, Oualid Abdelkader Bellag, Adibah Parman, Ibrahim Ali Noorbatcha, Mohd Noor Mat Isa, Muhammad Alfatih Muddathir Abdelrahim, et al.
    Sains Malaysiana, 2018;47:2951-2960.
    Metagenomic DNA library from palm oil mill effluent (POME) was constructed and subjected to high-throughput screening
    to find genes encoding cellulose- and xylan-degrading enzymes. DNA of 30 positive fosmid clones were sequenced with next
    generation sequencing technology and the raw data (short insert-paired) was analyzed with bioinformatic tools. First,
    the quality of 64,821,599 reverse and forward sequences of 101 bp length raw data was tested using Fastqc and SOLEXA.
    Then, raw data filtering was carried out by trimming low quality values and short reads and the vector sequences were
    removed and again the output was checked and the trimming was repeated until a high quality read sets was obtained.
    The second step was the de novo assembly of sequences to reconstruct 2900 contigs following de Bruijn graph algorithm.
    Pre-assembled contigs were arranged in order, the distances between contigs were identified and oriented with SSPACE,
    where 2139 scaffolds have been reconstructed. 16,386 genes have been identified after gene prediction using Prodigal
    and putative ID assignment with Blastp vs NR protein. The acceptable strategy to handle metagenomic NGS-data in order
    to detect known and potentially unknown genes is presented and we showed the computational efficiency of de Bruijn
    graph algorithm of de novo assembly to 21 bioprospect genes encoding cellulose-degrading enzymes and 6 genes
    encoding xylan-degrading enzymes of 30.3% to 100% identity percentage.
    Matched MeSH terms: Gene Library
  7. Fazia Adyani Ahmad Fuad, Houston Douglas R, Michels Paul AM, Fothergill-gilmore Linda A, Walkinshaw Malcolm D
    Sains Malaysiana, 2016;45:1113-1120.
    Cofactor-independent phosphoglycerate mutase has been proposed as a therapeutic target for the treatment of
    trypanosomatid diseases. In this paper, we report the identification of compounds that could potentially be developed as
    selective inhibitors of cofactor-independent phosphoglycerate mutase from Leishmania mexicana (LmiPGAM). Virtual
    screening was used in this search, as well as compounds identified by high-throughput screening. A ligand-based virtual
    screen programme, ultra fast shape recognition with atom types (UFSRAT), was used to screen for compounds resembling
    the substrate/product, before a structure-based approach was applied using AutoDock 4 and AutoDock Vina in a consensus
    docking scheme. In this way eight selected compounds were identified. In addition, three compounds from the Library of
    Pharmacologically Active Compounds (LOPAC) were selected from the published results of high-throughput screening of
    this library. The inhibitory effects of these compounds were tested at a fixed concentration of 1 mM. The results showed
    that seven compounds inhibited LmiPGAM activity and of these, two compounds (one each from high-throughput and
    virtual screening) showed substantial inhibition (i.e. 14% and 49% remaining activity, respectively). Taken together, the
    findings from this study indicate that these compounds have potential as novel inhibitors that specifically target LmiPGAM.
    Matched MeSH terms: Gene Library
  8. Wan KL, Chong SP, Ng ST, Shirley MW, Tomley FM, Jangi MS
    Int. J. Parasitol., 1999 Dec;29(12):1885-92.
    PMID: 10961844
    A study of about 500 expressed sequence tags (ESTs), derived from a merozoite cDNA library, was initiated as an approach to generate a larger pool of gene information on Eimeria tenella. Of the ESTs, 47.7% had matches with entries in the databases, including ribosomal proteins, metabolic enzymes and proteins with other functions, of which 14.3% represented previously known E. tenella genes. Thus over 50% of the ESTs had no significant database matches. The E. tenella EST dataset contained a range of highly abundant genes comparable with that found in the EST dataset of T. gondii and may thus reflect the importance of such molecules in the biology of the apicomplexan organisms. However, comparison of the two datasets revealed very few homologies between sequences of apical organelle molecules, and provides evidence for sequence divergence between these closely-related parasites. The data presented underpin the potential value of the EST strategy for the discovery of novel genes and may allow for a more rapid increase in the knowledge and understanding of gene expression in the merozoite life cycle stage of Eimeria spp.
    Matched MeSH terms: Gene Library
  9. Wong YC, Abd El Ghany M, Naeem R, Lee KW, Tan YC, Pain A, et al.
    Front Microbiol, 2016;7:1288.
    PMID: 27597847 DOI: 10.3389/fmicb.2016.01288
    Burkholderia cenocepacia infection often leads to fatal cepacia syndrome in cystic fibrosis patients. However, antibiotic therapy rarely results in complete eradication of the pathogen due to its intrinsic resistance to many clinically available antibiotics. Recent attention has turned to the identification of essential genes as the proteins encoded by these genes may serve as potential targets for development of novel antimicrobials. In this study, we utilized TraDIS (Transposon Directed Insertion-site Sequencing) as a genome-wide screening tool to facilitate the identification of B. cenocepacia genes essential for its growth and viability. A transposon mutant pool consisting of approximately 500,000 mutants was successfully constructed, with more than 400,000 unique transposon insertion sites identified by computational analysis of TraDIS datasets. The saturated library allowed for the identification of 383 genes that were predicted to be essential in B. cenocepacia. We extended the application of TraDIS to identify conditionally essential genes required for in vitro growth and revealed an additional repertoire of 439 genes to be crucial for B. cenocepacia growth under nutrient-depleted conditions. The library of B. cenocepacia mutants can subsequently be subjected to various biologically related conditions to facilitate the discovery of genes involved in niche adaptation as well as pathogenicity and virulence.
    Matched MeSH terms: Gene Library
  10. Loke KK, Rahnamaie-Tajadod R, Yeoh CC, Goh HH, Mohamed-Hussein ZA, Zainal Z, et al.
    PeerJ, 2017;5:e2938.
    PMID: 28265493 DOI: 10.7717/peerj.2938
    BACKGROUND: Polygonum minus is an herbal plant in the Polygonaceae family which is rich in ethnomedicinal plants. The chemical composition and characteristic pungent fragrance of Polygonum minus have been extensively studied due to its culinary and medicinal properties. There are only a few transcriptome sequences available for species from this important family of medicinal plants. The limited genetic information from the public expressed sequences tag (EST) library hinders further study on molecular mechanisms underlying secondary metabolite production.

    METHODS: In this study, we performed a hybrid assembly of 454 and Illumina sequencing reads from Polygonum minus root and leaf tissues, respectively, to generate a combined transcriptome library as a reference.

    RESULTS: A total of 34.37 million filtered and normalized reads were assembled into 188,735 transcripts with a total length of 136.67 Mbp. We performed a similarity search against all the publicly available genome sequences and found similarity matches for 163,200 (86.5%) of Polygonum minus transcripts, largely from Arabidopsis thaliana (58.9%). Transcript abundance in the leaf and root tissues were estimated and validated through RT-qPCR of seven selected transcripts involved in the biosynthesis of phenylpropanoids and flavonoids. All the transcripts were annotated against KEGG pathways to profile transcripts related to the biosynthesis of secondary metabolites.

    DISCUSSION: This comprehensive transcriptome profile will serve as a useful sequence resource for molecular genetics and evolutionary research on secondary metabolite biosynthesis in Polygonaceae family. Transcriptome assembly of Polygonum minus can be accessed at http://prims.researchfrontier.org/index.php/dataset/transcriptome.

    Matched MeSH terms: Gene Library
  11. Rodrigues, K. F., Tam, H. K.
    This paper describes the first reported attempt to isolate DNA sequences containing repeat motifs in Eurycoma longifolia and Orthosiphon stamineus. A library enriched for genomic repeat motifs was developed using novel oligonucleotides designed with inosine residues incorporated at predetermined positions. A total of eight and twelve specific molecular markers were developed for O. stamineus and E. longifolia respectively. These markers have a potential application in estimating population diversity levels and QTL mapping in these two medicinal plants, which are widely used in the Malaysian herbal industry.
    Matched MeSH terms: Gene Library
  12. Lee PS, Gan HM, Clements GR, Wilson JJ
    Genome, 2016 May 11.
    PMID: 27696907
    Mammal diversity assessments based on DNA derived from invertebrates have been suggested as alternatives to assessments based on traditional methods; however, no study has field-tested both approaches simultaneously. In Peninsular Malaysia, we calibrated the performance of mammal DNA derived from blowflies (Diptera: Calliphoridae) against traditional methods used to detect species. We first compared five methods (cage trapping, mist netting, hair trapping, scat collection, and blowfly-derived DNA) in a forest reserve with no recent reports of megafauna. Blowfly-derived DNA and mist netting detected the joint highest number of species (n = 6). Only one species was detected by multiple methods. Compared to the other methods, blowfly-derived DNA detected both volant and non-volant species. In another forest reserve, rich in megafauna, we calibrated blowfly-derived DNA against camera traps. Blowfly-derived DNA detected more species (n = 11) than camera traps (n = 9), with only one species detected by both methods. The rarefaction curve indicated that blowfly-derived DNA would continue to detect more species with greater sampling effort. With further calibration, blowfly-derived DNA may join the list of traditional field methods. Areas for further investigation include blowfly feeding and dispersal biology, primer biases, and the assembly of a comprehensive and taxonomically-consistent DNA barcode reference library.
    Matched MeSH terms: Gene Library
  13. Samad AFA, Sajad M, Jani J, Murad AMA, Ismail I
    Data Brief, 2018 Oct;20:555-557.
    PMID: 30197911 DOI: 10.1016/j.dib.2018.08.034
    Degradome sequencing referred as parallel analysis of RNA ends (PARE) by modifying 5'-rapid amplification of cDNA ends (RACE) with deep sequencing method. Deep sequencing of 5' products allow the determination of cleavage sites through the mapping of degradome fragments against small RNAs (miRNA or siRNA) on a large scale. Here, we carried out degradome sequencing in medicinal plant, Persicaria minor, to identify cleavage sites in small RNA libraries in control (mock-inoculated) and Fusarium oxysporum treated plants. The degradome library consisted of both control and treated samples which were pooled together during library preparation and named as D4. The D4 dataset have been deposited at GenBank under accession number SRX3921398, https://www.ncbi.nlm.nih.gov/sra/SRX3921398.
    Matched MeSH terms: Gene Library
  14. Teh KY, Afifudeen CLW, Aziz A, Wong LL, Loh SH, Cha TS
    Data Brief, 2019 Dec;27:104680.
    PMID: 31720332 DOI: 10.1016/j.dib.2019.104680
    Interest in harvesting potential benefits from microalgae renders it necessary to have the many ecological niches of a single species to be investigated. This dataset comprises de novo whole genome assembly of two mangrove-isolated microalgae (from division Chlorophyta); Chlorella vulgaris UMT-M1 and Messastrum gracile SE-MC4 from Universiti Malaysia Terengganu, Malaysia. Library runs were carried out with 2 × 150 base paired-ends reads, whereas sequencing was conducted using Illumina Novaseq 2500 platform. Sequencing yielded raw reads amounting to ∼11 Gb in total bases for both species and was further assembled de novo. Genome assembly resulted in a 50.15 Mbp and 60.83 Mbp genome size for UMT-M1 and SE-MC4, respectively. All filtered and assembled genomic data sequences have been submitted to National Centre for Biotechnology Information (NCBI) and can be located at DDBJ/ENA/GenBank under the accession of VJNP00000000 (UMT-M1) and VIYE00000000 (SE-MC4).
    Matched MeSH terms: Gene Library
  15. Nadiah Abu, Noraini Nordin, Noorjahan Banu Alitheen, Nadiah Abu, Sheau Wei Tan, Swee Keong Yeap, et al.
    Sains Malaysiana, 2018;47:303-308.
    RNA-seq has become an essential tool in molecular research. Nevertheless, application of RNA-seq was limited by cost and technical difficulties. Illumina has introduced the cost effective and ease to handle Truseq Targeted RNA Sequencing. In this study, we present the requirements and the optimization procedure for this Truseq Targeted RNA sequencing on cell line. Total RNA was recommended as starting materials but it required optimization including additional purification step and adjusting the AMPure beads ratio to eliminate unwanted contaminants. This can be resolved by using PolyA-enriched mRNA as starting material. TREx is a useful assay to evaluate gene expression. Quality library of TREx can be prepared by adding multiple washing steps or changing input sample to mRNA.
    Matched MeSH terms: Gene Library
  16. Fadli N, Mohd Nor SA, Othman AS, Sofyan H, Muchlisin ZA
    PeerJ, 2020;8:e9641.
    PMID: 32844060 DOI: 10.7717/peerj.9641
    Knowledge on the precise identification of fish resources is critical for sustainable fisheries management. This study employs the DNA barcoding approach to generate a molecular taxonomic catalogue of commercially important reef fishes in the waters of Weh Island (Aceh Province), the most northerly inhabited island in the biodiverse Indonesian Archipelago. The waters not only support artisanal fisheries but also a feeder for the industry in the greater island of Aceh. In total, 230 specimens from 72 species belonging to 32 genera and 17 families were DNA barcoded, representing a major segment of the captured reef fish taxa and a quarter of fish species diversity that had previously been recorded. The sequence read lengths were 639 bp revealing 359 conserved sites, 280 variable sites, 269 parsimony informative and 11 singletons. Our molecular findings paralleled the morphological identification with no evidence of cryptic species or new species discovery. This study is a significant contribution to the fisheries statistics of this area, which would facilitate assessment of species catch composition and hence for strategizing management plans. It is an important input to the DNA barcode library of Indonesian marine fishes and to the global DNA barcode entries in general.
    Matched MeSH terms: Gene Library
  17. Yew CW, Kumar SV
    Mol Biol Rep, 2012 Feb;39(2):1783-90.
    PMID: 21625851 DOI: 10.1007/s11033-011-0919-7
    MicroRNAs (miRNAs) are small RNAs (sRNAs) with approximately 21-24 nucleotides in length. They regulate the expression of target genes through the mechanism of RNA silencing. Conventional isolation and cloning of miRNAs methods are usually technical demanding and inefficient. These limitations include the requirement for high amounts of starting total RNA, inefficient ligation of linkers, high amount of PCR artifacts and bias in the formation of short miRNA-concatamers. Here we describe in detail a method that uses 80 μg of total RNA as the starting material. Enhancement of the ligation of sRNAs and linkers with the use of polyethylene glycol (PEG8000) was described. PCR artifacts from the amplification of reverse-transcribed sRNAs were greatly decreased by using lower concentrations of primers and reducing the number of amplification cycles. Large concatamers with up to 1 kb in size with around 20 sRNAs/concatamer were obtained by using an optimized reaction condition. This protocol provide researchers with a rapid, efficient and cost-effective method for the construction of miRNA profiles from plant tissues containing low amounts of total RNA, such as fruit flesh and senescent leaves.
    Matched MeSH terms: Gene Library*
  18. Lim BN, Choong YS, Ismail A, Glökler J, Konthur Z, Lim TS
    Biotechniques, 2012 Dec;53(6):357-64.
    PMID: 23227986 DOI: 10.2144/000113964
    Directed evolution of nucleotide libraries using recombination or mutagenesis is an important technique for customizing catalytic or biophysical traits of proteins. Conventional directed evolution methods, however, suffer from cumbersome digestion and ligation steps. Here, we describe a simple method to increase nucleotide diversity using single-stranded DNA (ssDNA) as a starting template. An initial PCR amplification using phosphorylated primers with overlapping regions followed by treatment with lambda exonuclease generates ssDNA templates that can then be annealed via the overlap regions. Double-stranded DNA (dsDNA) is then generated through extension with Klenow fragment. To demonstrate the applicability of this methodology for directed evolution of nucleotide libraries, we generated both gene shuffled and regional mutagenesis synthetic antibody libraries with titers of 2×108 and 6×107, respectively. We conclude that our method is an efficient and convenient approach to generate diversity in nucleic acid based libraries, especially recombinant antibody libraries.
    Matched MeSH terms: Gene Library*
  19. Ng CY, Wickneswari R, Choong CY
    Genet. Mol. Res., 2014;13(3):6037-49.
    PMID: 25117361 DOI: 10.4238/2014.August.7.18
    Calamus palustris Griff. is an economically important dioecious rattan species in Southeast Asia. However, dioecy and onset of flowering at 3-4 years old render uncertainties in desired female:male seedling ratios to establish a productive seed orchard for this rattan species. We constructed a subtractive library for male floral tissue to understand the genetic mechanism for gender determination in C. palustris. The subtractive library produced 1536 clones with 1419 clones of high quality. Reverse Northern screening showed 313 clones with differential expression, and sequence analyses clustered them into 205 unigenes, including 32 contigs and 173 singletons. The subtractive library was further validated with reverse transcription-quantitative polymerase chain reaction analysis. Homology identification classified the unigenes into 12 putative functional proteins with 83% unigenes showing significant match to proteins in databases. Functional annotations of these unigenes revealed genes involved in male flower development, including MADS-box genes, pollen-related genes, phytohormones for flower development, and male flower organ development. Our results showed that the male floral genes may play a vital role in sex determination in C. palustris. The identified genes can be exploited to understand the molecular basis of sex determination in C. palustris.
    Matched MeSH terms: Gene Library
  20. Ho WS, Pang SL, Abdullah J
    Physiol Mol Biol Plants, 2014 Jul;20(3):393-7.
    PMID: 25049467 DOI: 10.1007/s12298-014-0230-x
    The large-scale genomic resource for kelampayan was generated from a developing xylem cDNA library. A total of 6,622 high quality expressed sequence tags (ESTs) were generated through high-throughput 5' EST sequencing of cDNA clones. The ESTs were analyzed and assembled to generate 4,728 xylogenesis unigenes distributed in 2,100 contigs and 2,628 singletons. About 59.3 % of the ESTs were assigned with putative identifications whereas 40.7 % of the sequences showed no significant similarity to any sequences in GenBank. Interestingly, most genes involved in lignin biosynthesis and several other cell wall biosynthesis genes were identified in the kelampayan EST database. The identified genes in this study will be candidates for functional genomics and association genetic studies in kelampayan aiming at the production of high value forests.
    Matched MeSH terms: Gene Library
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