Displaying publications 1 - 20 of 22 in total

Abstract:
Sort:
  1. A Review of Gene Knockout Strategies for Microbial Cells
    Tang PW, Chua PS, Chong SK, Mohamad MS, Choon YW, Deris S, et al.
    Recent Pat Biotechnol, 2015;9(3):176-97.
    PMID: 27185502
    BACKGROUND: Predicting the effects of genetic modification is difficult due to the complexity of metabolic net- works. Various gene knockout strategies have been utilised to deactivate specific genes in order to determine the effects of these genes on the function of microbes. Deactivation of genes can lead to deletion of certain proteins and functions. Through these strategies, the associated function of a deleted gene can be identified from the metabolic networks.

    METHODS: The main aim of this paper is to review the available techniques in gene knockout strategies for microbial cells. The review is done in terms of their methodology, recent applications in microbial cells. In addition, the advantages and disadvantages of the techniques are compared and discuss and the related patents are also listed as well.

    RESULTS: Traditionally, gene knockout is done through wet lab (in vivo) techniques, which were conducted through laboratory experiments. However, these techniques are costly and time consuming. Hence, various dry lab (in silico) techniques, where are conducted using computational approaches, have been developed to surmount these problem.

    CONCLUSION: The development of numerous techniques for gene knockout in microbial cells has brought many advancements in the study of gene functions. Based on the literatures, we found that the gene knockout strategies currently used are sensibly implemented with regard to their benefits.

    Matched MeSH terms: Gene Knockout Techniques/methods*
  2. Identification of gene knockout strategies using a hybrid of an ant colony optimization algorithm and flux balance analysis to optimize microbial strains
    Lu SJ, Salleh AH, Mohamad MS, Deris S, Omatu S, Yoshioka M
    Comput Biol Chem, 2014 12;53PB:175-183.
    PMID: 25462325 DOI: 10.1016/j.compbiolchem.2014.09.008
    Reconstructions of genome-scale metabolic networks from different organisms have become popular in recent years. Metabolic engineering can simulate the reconstruction process to obtain desirable phenotypes. In previous studies, optimization algorithms have been implemented to identify the near-optimal sets of knockout genes for improving metabolite production. However, previous works contained premature convergence and the stop criteria were not clear for each case. Therefore, this study proposes an algorithm that is a hybrid of the ant colony optimization algorithm and flux balance analysis (ACOFBA) to predict near optimal sets of gene knockouts in an effort to maximize growth rates and the production of certain metabolites. Here, we present a case study that uses Baker's yeast, also known as Saccharomyces cerevisiae, as the model organism and target the rate of vanillin production for optimization. The results of this study are the growth rate of the model organism after gene deletion and a list of knockout genes. The ACOFBA algorithm was found to improve the yield of vanillin in terms of growth rate and production compared with the previous algorithms.
    Matched MeSH terms: Gene Knockout Techniques
  3. Fulltext In vivo assembly of DNA-fragments in the moss, Physcomitrella patens
    King BC, Vavitsas K, Ikram NK, Schrøder J, Scharff LB, Bassard JÉ, et al.
    Sci Rep, 2016 04 29;6:25030.
    PMID: 27126800 DOI: 10.1038/srep25030
    Direct assembly of multiple linear DNA fragments via homologous recombination, a phenomenon known as in vivo assembly or transformation associated recombination, is used in biotechnology to assemble DNA constructs ranging in size from a few kilobases to full synthetic microbial genomes. It has also enabled the complete replacement of eukaryotic chromosomes with heterologous DNA. The moss Physcomitrella patens, a non-vascular and spore producing land plant (Bryophyte), has a well-established capacity for homologous recombination. Here, we demonstrate the in vivo assembly of multiple DNA fragments in P. patens with three examples of effective genome editing: we (i) efficiently deleted a genomic locus for diterpenoid metabolism yielding a biosynthetic knockout, (ii) introduced a salt inducible promoter, and (iii) re-routed endogenous metabolism into the formation of amorphadiene, a precursor of high-value therapeutics. These proof-of-principle experiments pave the way for more complex and increasingly flexible approaches for large-scale metabolic engineering in plant biotechnology.
    Matched MeSH terms: Gene Knockout Techniques
  4. A hybrid of Cuckoo Search and Minimization of Metabolic Adjustment to optimize metabolites production in genome-scale models
    Arif MA, Mohamad MS, Abd Latif MS, Deris S, Remli MA, Mohd Daud K, et al.
    Comput Biol Med, 2018 11 01;102:112-119.
    PMID: 30267898 DOI: 10.1016/j.compbiomed.2018.09.015
    Metabolic engineering involves the modification and alteration of metabolic pathways to improve the production of desired substance. The modification can be made using in silico gene knockout simulation that is able to predict and analyse the disrupted genes which may enhance the metabolites production. Global optimization algorithms have been widely used for identifying gene knockout strategies. However, their productions were less than theoretical maximum and the algorithms are easily trapped into local optima. These algorithms also require a very large computation time to obtain acceptable results. This is due to the complexity of the metabolic models which are high dimensional and contain thousands of reactions. In this paper, a hybrid algorithm of Cuckoo Search and Minimization of Metabolic Adjustment is proposed to overcome the aforementioned problems. The hybrid algorithm searches for the near-optimal set of gene knockouts that leads to the overproduction of metabolites. Computational experiments on two sets of genome-scale metabolic models demonstrate that the proposed algorithm is better than the previous works in terms of growth rate, Biomass Product Couple Yield, and computation time.
    Matched MeSH terms: Gene Knockout Techniques
  5. Fulltext In silico gene knockout prediction using a hybrid of Bat algorithm and minimization of metabolic adjustment
    Man MY, Mohamad MS, Choon YW, Ismail MA
    J Integr Bioinform, 2021 Aug 04;18(3).
    PMID: 34348418 DOI: 10.1515/jib-2020-0037
    Microorganisms commonly produce many high-demand industrial products like fuels, food, vitamins, and other chemicals. Microbial strains are the strains of microorganisms, which can be optimized to improve their technological properties through metabolic engineering. Metabolic engineering is the process of overcoming cellular regulation in order to achieve a desired product or to generate a new product that the host cells do not usually need to produce. The prediction of genetic manipulations such as gene knockout is part of metabolic engineering. Gene knockout can be used to optimize the microbial strains, such as to maximize the production rate of chemicals of interest. Metabolic and genetic engineering is important in producing the chemicals of interest as, without them, the product yields of many microorganisms are normally low. As a result, the aim of this paper is to propose a combination of the Bat algorithm and the minimization of metabolic adjustment (BATMOMA) to predict which genes to knock out in order to increase the succinate and lactate production rates in Escherichia coli (E. coli).
    Matched MeSH terms: Gene Knockout Techniques
  6. Fulltext Differential Bees Flux Balance Analysis with OptKnock for in silico microbial strains optimization
    Choon YW, Mohamad MS, Deris S, Illias RM, Chong CK, Chai LE, et al.
    PLoS One, 2014;9(7):e102744.
    PMID: 25047076 DOI: 10.1371/journal.pone.0102744
    Microbial strains optimization for the overproduction of desired phenotype has been a popular topic in recent years. The strains can be optimized through several techniques in the field of genetic engineering. Gene knockout is a genetic engineering technique that can engineer the metabolism of microbial cells with the objective to obtain desirable phenotypes. However, the complexities of the metabolic networks have made the process to identify the effects of genetic modification on the desirable phenotypes challenging. Furthermore, a vast number of reactions in cellular metabolism often lead to the combinatorial problem in obtaining optimal gene deletion strategy. Basically, the size of a genome-scale metabolic model is usually large. As the size of the problem increases, the computation time increases exponentially. In this paper, we propose Differential Bees Flux Balance Analysis (DBFBA) with OptKnock to identify optimal gene knockout strategies for maximizing the production yield of desired phenotypes while sustaining the growth rate. This proposed method functions by improving the performance of a hybrid of Bees Algorithm and Flux Balance Analysis (BAFBA) by hybridizing Differential Evolution (DE) algorithm into neighborhood searching strategy of BAFBA. In addition, DBFBA is integrated with OptKnock to validate the results for improving the reliability the work. Through several experiments conducted on Escherichia coli, Bacillus subtilis, and Clostridium thermocellum as the model organisms, DBFBA has shown a better performance in terms of computational time, stability, growth rate, and production yield of desired phenotypes compared to the methods used in previous works.
    Matched MeSH terms: Gene Knockout Techniques/methods*
  7. In silico deletion of PtsG gene in Escherichia coli genome-scale model predicts increased succinate production from glycerol
    Mienda BS, Shamsir MS
    J Biomol Struct Dyn, 2015;33(11):2380-9.
    PMID: 25921851 DOI: 10.1080/07391102.2015.1036461
    Systems metabolic engineering and in silico analyses are necessary to study gene knockout candidate for enhanced succinic acid production by Escherichia coli. Metabolically engineered E. coli has been reported to produce succinate from glucose and glycerol. However, investigation on in silico deletion of ptsG/b1101 gene in E. coli from glycerol using minimization of metabolic adjustment algorithm with the OptFlux software platform has not yet been elucidated. Herein we report what is to our knowledge the first direct predicted increase in succinate production following in silico deletion of the ptsG gene in E. coli GEM from glycerol with the OptFlux software platform. The result indicates that the deletion of this gene in E. coli GEM predicts increased succinate production that is 20% higher than the wild-type control model. Hence, the mutant model maintained a growth rate that is 77% of the wild-type parent model. It was established that knocking out of the ptsG/b1101 gene in E. coli using glucose as substrate enhanced succinate production, but the exact mechanism of this effect is still obscure. This study informs other studies that the deletion of ptsG/b1101 gene in E. coli GEM predicted increased succinate production, enabling a model-driven experimental inquiry and/or novel biological discovery on the underground metabolic role of this gene in E. coli central metabolism in relation to increasing succinate production when glycerol is the substrate.
    Matched MeSH terms: Gene Knockout Techniques
  8. Fulltext Targeted inactivation of Salmonella Agona metabolic genes by group II introns and in vivo assessment of pathogenicity and anti-tumour activity in mouse model
    Gwee CP, Khoo CH, Yeap SK, Tan GC, Cheah YK
    PeerJ, 2019;7:e5989.
    PMID: 30671294 DOI: 10.7717/peerj.5989
    The fight against cancer has been a never-ending battle. Limitations of conventional therapies include lack of selectivity, poor penetration and highly toxic to the host. Using genetically modified bacteria as a tumour therapy agent has gained the interest of scientist from the past few decades. Low virulence and highly tolerability of Salmonella spp. in animals and humans make it as the most studied pathogen with regards to anti-tumour therapy. The present study aims to construct a genetically modified S. Agona auxotroph as an anti-tumour agent. LeuB and ArgD metabolic genes in ΔSopBΔSopD double knockout S. Agona were successfully knocked out using a Targetron gene knockout system. The knockout was confirmed by colony PCR and the strains were characterized in vitro and in vivo. The knockout of metabolic genes causes significant growth defect in M9 minimal media. Quadruple knockout ΔSopBΔSopDΔLeuBΔArgD (BDLA) exhibited lowest virulence among all of the strains in all parameters including bacterial load, immunity profile and histopathology studies. In vivo anti-tumour study on colorectal tumour bearing-BALB/c mice revealed that all strains of S. Agona were able to suppress the growth of the large solid tumour as compared with negative control and ΔLeuBΔArgD (LA) and BDLA auxotroph showed better efficacy. Interestingly, higher level of tumour growth suppression was noticed in large tumour. However, multiple administration of bacteria dosage did not increase the tumour suppression efficacy. In this study, the virulence of BDLA knockout strain was slightly reduced and tumour growth suppression efficacy was successfully enhanced, which provide a valuable starting point for the development of S. Agona as anti-tumour agent.
    Matched MeSH terms: Gene Knockout Techniques
  9. Model-aided atpE gene knockout strategy in Escherichia coli for enhanced succinic acid production from glycerol
    Mienda BS, Shamsir MS, Md Illias R
    J Biomol Struct Dyn, 2016 Aug;34(8):1705-16.
    PMID: 26513379 DOI: 10.1080/07391102.2015.1090341
    Succinic acid is an important platform chemical with a variety of applications. Model-guided metabolic engineering strategies in Escherichia coli for strain improvement to increase succinic acid production using glucose and glycerol remain largely unexplored. Herein, we report what are, to our knowledge, the first metabolic knockout of the atpE gene to have increased succinic acid production using both glucose and alternative glycerol carbon sources in E. coli. Guided by a genome-scale metabolic model, we engineered the E. coli host to enhance anaerobic production of succinic acid by deleting the atpE gene, thereby generating additional reducing equivalents by blocking H(+) conduction across the mutant cell membrane. This strategy produced 1.58 and .49 g l(-1) of succinic acid from glycerol and glucose substrate, respectively. This work further elucidates a model-guided and/or system-based metabolic engineering, involving only a single-gene deletion strategy for enhanced succinic acid production in E. coli.
    Matched MeSH terms: Gene Knockout Techniques*
  10. Fulltext Deep Brain Photoreceptor (val-opsin) Gene Knockout Using CRISPR/Cas Affects Chorion Formation and Embryonic Hatching in the Zebrafish
    Hang CY, Moriya S, Ogawa S, Parhar IS
    PLoS One, 2016;11(10):e0165535.
    PMID: 27792783 DOI: 10.1371/journal.pone.0165535
    Non-rod non-cone photopigments in the eyes and the brain can directly mediate non-visual functions of light in non-mammals. This was supported by our recent findings on vertebrate ancient long (VAL)-opsin photopigments encoded by the val-opsinA (valopa) and val-opsinB (valopb) genes in zebrafish. However, the physiological functions of valop isoforms remain unknown. Here, we generated valop-mutant zebrafish using CRISPR/Cas genome editing, and examined the phenotypes of loss-of-function mutants. F0 mosaic mutations and germline transmission were confirmed via targeted insertions and/or deletions in the valopa or valopb gene in F1 mutants. Based on in silico analysis, frameshift mutations converted VAL-opsin proteins to non-functional truncated forms with pre-mature stop codons. Most F1 eggs or embryos from F0 female valopa/b mutants showed either no or only partial chorion elevation, and the eggs or embryos died within 26 hour-post-fertilization. However, most F1 embryos from F0 male valopa mutant developed but hatched late compared to wild-type embryos, which hatched at 4 day-post-fertilization. Late-hatched F1 offspring included wild-type and mutants, indicating the parental effects of valop knockout. This study shows valop gene knockout affects chorion formation and embryonic hatching in the zebrafish.
    Matched MeSH terms: Gene Knockout Techniques*
  11. A hybrid of ant colony optimization and minimization of metabolic adjustment to improve the production of succinic acid in Escherichia coli
    Chong SK, Mohamad MS, Mohamed Salleh AH, Choon YW, Chong CK, Deris S
    Comput Biol Med, 2014 Jun;49:74-82.
    PMID: 24763079 DOI: 10.1016/j.compbiomed.2014.03.011
    This paper presents a study on gene knockout strategies to identify candidate genes to be knocked out for improving the production of succinic acid in Escherichia coli. Succinic acid is widely used as a precursor for many chemicals, for example production of antibiotics, therapeutic proteins and food. However, the chemical syntheses of succinic acid using the traditional methods usually result in the production that is far below their theoretical maximums. In silico gene knockout strategies are commonly implemented to delete the gene in E. coli to overcome this problem. In this paper, a hybrid of Ant Colony Optimization (ACO) and Minimization of Metabolic Adjustment (MoMA) is proposed to identify gene knockout strategies to improve the production of succinic acid in E. coli. As a result, the hybrid algorithm generated a list of knockout genes, succinic acid production rate and growth rate for E. coli after gene knockout. The results of the hybrid algorithm were compared with the previous methods, OptKnock and MOMAKnock. It was found that the hybrid algorithm performed better than OptKnock and MOMAKnock in terms of the production rate. The information from the results produced from the hybrid algorithm can be used in wet laboratory experiments to increase the production of succinic acid in E. coli.
    Matched MeSH terms: Gene Knockout Techniques
  12. Fulltext NDV-induced apoptosis in absence of Bax; evidence of involvement of apoptotic proteins upstream of mitochondria
    Molouki A, Yusoff K
    Virol J, 2012;9:179.
    PMID: 22935147 DOI: 10.1186/1743-422X-9-179
    Recently it was shown that following infection of HeLa cells with Newcastle disease virus (NDV), the matrix (M) protein binds to Bax and subsequently the intrinsic pathway of apoptosis is activated. Moreover, there was very little alteration on mRNA and protein levels of Bax and Bcl-2 after infection with NDV.
    Matched MeSH terms: Gene Knockout Techniques
  13. Model-assisted formate dehydrogenase-O (fdoH) gene knockout for enhanced succinate production in Escherichia coli from glucose and glycerol carbon sources
    Mienda BS, Shamsir MS, Md Illias R
    J Biomol Struct Dyn, 2016 Nov;34(11):2305-16.
    PMID: 26510527 DOI: 10.1080/07391102.2015.1113387
    Succinic acid is an important platform chemical that has broad applications and is been listed as one of the top twelve bio-based chemicals produced from biomass by the US Department of Energy. The metabolic role of Escherichia coli formate dehydrogenase-O (fdoH) under anaerobic conditions in relation to succinic acid production remained largely unspecified. Herein we report, what are to our knowledge, the first metabolic fdoH gene knockout that have enhanced succinate production using glucose and glycerol substrates in E. coli. Using the most recent E. coli reconstruction iJO1366, we engineered its host metabolism to enhance the anaerobic succinate production by deleting the fdoH gene, which blocked H(+) conduction across the mutant cell membrane for the enhanced succinate production. The engineered mutant strain BMS4 showed succinate production of 2.05 g l(-1) (41.2-fold in 7 days) from glycerol and .39 g l(-1) (6.2-fold in 1 day) from glucose. This work revealed that a single deletion of the fdoH gene is sufficient to increase succinate production in E. coli from both glucose and glycerol substrates.
    Matched MeSH terms: Gene Knockout Techniques
  14. Model-guided metabolic gene knockout of gnd for enhanced succinate production in Escherichia coli from glucose and glycerol substrates
    Mienda BS, Shamsir MS, Illias RM
    Comput Biol Chem, 2016 Apr;61:130-7.
    PMID: 26878126 DOI: 10.1016/j.compbiolchem.2016.01.013
    The metabolic role of 6-phosphogluconate dehydrogenase (gnd) under anaerobic conditions with respect to succinate production in Escherichia coli remained largely unspecified. Herein we report what are to our knowledge the first metabolic gene knockout of gnd to have increased succinic acid production using both glucose and glycerol substrates in E. coli. Guided by a genome scale metabolic model, we engineered the E. coli host metabolism to enhance anaerobic production of succinic acid by deleting the gnd gene, considering its location in the boundary of oxidative and non-oxidative pentose phosphate pathway. This strategy induced either the activation of malic enzyme, causing up-regulation of phosphoenolpyruvate carboxylase (ppc) and down regulation of phosphoenolpyruvate carboxykinase (ppck) and/or prevents the decarboxylation of 6 phosphogluconate to increase the pool of glyceraldehyde-3-phosphate (GAP) that is required for the formation of phosphoenolpyruvate (PEP). This approach produced a mutant strain BMS2 with succinic acid production titers of 0.35gl(-1) and 1.40gl(-1) from glucose and glycerol substrates respectively. This work further clearly elucidates and informs other studies that the gnd gene, is a novel deletion target for increasing succinate production in E. coli under anaerobic condition using glucose and glycerol carbon sources. The knowledge gained in this study would help in E. coli and other microbial strains development for increasing succinate production and/or other industrial chemicals.
    Matched MeSH terms: Gene Knockout Techniques
  15. Fulltext Analysis of the xplAB-containing gene cluster involved in the bacterial degradation of the explosive hexahydro-1,3,5-trinitro-1,3,5-triazine
    Chong CS, Sabir DK, Lorenz A, Bontemps C, Andeer P, Stahl DA, et al.
    Appl Environ Microbiol, 2014 Nov;80(21):6601-10.
    PMID: 25128343 DOI: 10.1128/AEM.01818-14
    Repeated use of the explosive compound hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) on military land has resulted in significant soil and groundwater pollution. Rates of degradation of RDX in the environment are low, and accumulated RDX, which the U.S. Environmental Protection Agency has determined is a possible human carcinogen, is now threatening drinking water supplies. RDX-degrading microorganisms have been isolated from RDX-contaminated land; however, despite the presence of these species in contaminated soils, RDX pollution persists. To further understand this problem, we studied RDX-degrading species belonging to four different genera (Rhodococcus, Microbacterium, Gordonia, and Williamsia) isolated from geographically distinct locations and established that the xplA and xplB (xplAB) genes, which encode a cytochrome P450 and a flavodoxin redox partner, respectively, are nearly identical in all these species. Together, the xplAB system catalyzes the reductive denitration of RDX and subsequent ring cleavage under aerobic and anaerobic conditions. In addition to xplAB, the Rhodococcus species studied here share a 14-kb region flanking xplAB; thus, it appears likely that the RDX-metabolizing ability was transferred as a genomic island within a transposable element. The conservation and transfer of xplAB-flanking genes suggest a role in RDX metabolism. We therefore independently knocked out genes within this cluster in the RDX-degrading species Rhodococcus rhodochrous 11Y. Analysis of the resulting mutants revealed that XplA is essential for RDX degradation and that XplB is not the sole contributor of reducing equivalents to XplA. While XplA expression is induced under nitrogen-limiting conditions and further enhanced by the presence of RDX, MarR is not regulated by RDX.
    Matched MeSH terms: Gene Knockout Techniques
  16. Optimising the production of succinate and lactate in Escherichia coli using a hybrid of artificial bee colony algorithm and minimisation of metabolic adjustment
    Tang PW, Choon YW, Mohamad MS, Deris S, Napis S
    J Biosci Bioeng, 2015 Mar;119(3):363-8.
    PMID: 25216804 DOI: 10.1016/j.jbiosc.2014.08.004
    Metabolic engineering is a research field that focuses on the design of models for metabolism, and uses computational procedures to suggest genetic manipulation. It aims to improve the yield of particular chemical or biochemical products. Several traditional metabolic engineering methods are commonly used to increase the production of a desired target, but the products are always far below their theoretical maximums. Using numeral optimisation algorithms to identify gene knockouts may stall at a local minimum in a multivariable function. This paper proposes a hybrid of the artificial bee colony (ABC) algorithm and the minimisation of metabolic adjustment (MOMA) to predict an optimal set of solutions in order to optimise the production rate of succinate and lactate. The dataset used in this work was from the iJO1366 Escherichia coli metabolic network. The experimental results include the production rate, growth rate and a list of knockout genes. From the comparative analysis, ABCMOMA produced better results compared to previous works, showing potential for solving genetic engineering problems.
    Matched MeSH terms: Gene Knockout Techniques
  17. Fulltext Identification of curcumin analogues with anti-seizure potential in vivo using chemical and genetic zebrafish larva seizure models
    Choo BKM, Kundap UP, Faudzi SMM, Abas F, Shaikh MF, Samarut É
    Biomed Pharmacother, 2021 Oct;142:112035.
    PMID: 34411917 DOI: 10.1016/j.biopha.2021.112035
    Seizures are the outward manifestation of abnormally excessive or synchronous brain activity. While seizures can be somewhat symptomatically managed with anti-epileptic drugs (AEDs), many patients are still refractory to the currently available AEDs. As a result, there is a need to identify new molecules with anti-seizure properties. Curcumin is the principle curcuminoid of Curcuma longa, or colloquially turmeric, and has been experimentally proven to have anti-convulsive properties, but its poor bioavailability has dampened further therapeutic interest. Hence, this study aimed to ask if structural analogues of curcumin with an adequate bioavailability could have an anti-seizure effect in vivo. To do so, we tested these analogues following a multipronged approach combining the use of several zebrafish seizure models (chemically-induced and genetic) and complementary assays (behavioural and brain activity). Overall, from the 68 analogues tested, we found 15 different derivatives that were able to significantly decrease the behavioural hyperactivity induced by pentylenetetrazol. Of those, only a few showed an effect on the hyperactivity phenotype of two genetic models of brain seizures that are the gabra1 and gabrg2 knockouts. Two analogues, CA 80(1) and CA 74(1), were able to significantly alleviate brain seizures of gabrg2-mutant larvae. As a result, these analogues are good candidates as novel anti-seizure agents.
    Matched MeSH terms: Gene Knockout Techniques
  18. Fulltext The kiss/kissr systems are dispensable for zebrafish reproduction: evidence from gene knockout studies
    Tang H, Liu Y, Luo D, Ogawa S, Yin Y, Li S, et al.
    Endocrinology, 2015 Feb;156(2):589-99.
    PMID: 25406015 DOI: 10.1210/en.2014-1204
    The kiss1/gpr54 signaling system is considered to be a critical regulator of reproduction in most vertebrates. However, this presumption has not been tested vigorously in nonmammalian vertebrates. Distinct from mammals, multiple kiss1/gpr54 paralogous genes (kiss/kissr) have been identified in nonmammalian vertebrates, raising the possibility of functional redundancy among these genes. In this study, we have systematically generated the zebrafish kiss1(-/-), kiss2(-/-), and kiss1(-/-);kiss2(-/-) mutant lines as well as the kissr1(-/-), kissr2(-/-), and kissr1(-/-);kissr2(-/-) mutant lines using transcription activator-like effector nucleases. We have demonstrated that spermatogenesis and folliculogenesis as well as reproductive capability are not impaired in all of these 6 mutant lines. Collectively, our results indicate that kiss/kissr signaling is not absolutely required for zebrafish reproduction, suggesting that the kiss/kissr systems play nonessential roles for reproduction in certain nonmammalian vertebrates. These findings also demonstrated that fish and mammals have evolved different strategies for neuroendocrine control of reproduction.
    Matched MeSH terms: Gene Knockout Techniques
  19. Pathogenicity and phenotypic analysis of sopB, sopD and pipD virulence factors in Salmonella enterica serovar typhimurium and Salmonella enterica serovar Agona
    Khoo CH, Sim JH, Salleh NA, Cheah YK
    Antonie Van Leeuwenhoek, 2015 Jan;107(1):23-37.
    PMID: 25312847 DOI: 10.1007/s10482-014-0300-7
    Salmonella is an important food-borne pathogen causing disease in humans and animals worldwide. Salmonellosis may be caused by any one of over 2,500 serovars of Salmonella. Nonetheless, Salmonella enterica serovar Typhimurium and Salmonella enterica serovar Agona are the second most prevalent serovars isolated from humans and livestock products respectively. Limited knowledge is available about the virulence mechanisms responsible for diarrheal disease caused by them. To investigate the contribution of sopB, sopD and pipD as virulence factors in intracellular infections and the uniqueness of these bacteria becoming far more prevalent than other serovars, the infection model of Caenorhabditis elegans and phenotypic microarray were used to characterize their mutants. The strains containing the mutation in sopB, sopD and pipD genes were constructed by using latest site-specific group II intron mutagenesis approach to reveal the pathogenicity of the virulence factors. Overall, we observed that the mutations in sopB, sopD and pipD genes of both serovars did not exhibit significant decrease in virulence towards the nematode. This may indicate that these virulence effectors may not be universal virulence factors involved in conserved innate immunity. There are significant phenotypic differences amongst strains carrying sopB, sopD and pipD gene mutations via the analysis of biochemical profiles of the bacteria. Interestingly, mutant strains displayed different susceptibility to chemical stressors from several distinct pharmacological and structural classes when compared to its isogenic parental strains. These metabolic and chemosensitivity assays also revealed multiple roles of Salmonella virulence factors in nutrient metabolism and antibiotic resistance.
    Matched MeSH terms: Gene Knockout Techniques
  20. Escherichia coli strains expressing H12 antigens demonstrate an increased ability to attach to abiotic surfaces as compared with E. coli strains expressing H7 antigens
    Goulter RM, Taran E, Gentle IR, Gobius KS, Dykes GA
    Colloids Surf B Biointerfaces, 2014 Jul 1;119:90-8.
    PMID: 24880987 DOI: 10.1016/j.colsurfb.2014.04.003
    The role of Escherichia coli H antigens in hydrophobicity and attachment to glass, Teflon and stainless steel (SS) surfaces was investigated through construction of fliC knockout mutants in E. coli O157:H7, O1:H7 and O157:H12. Loss of FliC(H12) in E. coli O157:H12 decreased attachment to glass, Teflon and stainless steel surfaces (p<0.05). Complementing E. coli O157:H12 ΔfliC(H12) with cloned wildtype (wt) fliC(H12) restored attachment to wt levels. The loss of FliCH7 in E. coli O157:H7 and O1:H7 did not always alter attachment (p>0.05), but complementation with cloned fliC(H12), as opposed to cloned fliCH7, significantly increased attachment for both strains compared with wt counterparts (p<0.05). Hydrophobicity determined using bacterial adherence to hydrocarbons and contact angle measurements differed with fliC expression but was not correlated to the attachment to materials included in this study. Purified FliC was used to functionalise silicone nitride atomic force microscopy probes, which were used to measure adhesion forces between FliC and substrates. Although no significant difference in adhesion force was observed between FliC(H12) and FliCH7 probes, differences in force curves suggest different mechanism of attachment for FliC(H12) compared with FliCH7. These results indicate that E. coli strains expressing flagellar H12 antigens have an increased ability to attach to certain abiotic surfaces compared with E. coli strains expressing H7 antigens.
    Matched MeSH terms: Gene Knockout Techniques
Related Terms
Filters
Contact Us

Please provide feedback to Administrator (afdal@afpm.org.my)

External Links