Journal, genetic engineering & biotechnology最新文献

Nowadays, breakthroughs in molecular biology are happening at an unprecedented rate. One of them is the ability to engineer transgenic animals. A transgenic animal is one whose genome has been changed to carry genes from another species or to use techniques for animal genome editing for specific traits. Animal features can be changed by purposefully altering the gene (or genes). A mouse was the first successful transgenic animal. Then pigs, sheep, cattle, and rabbits came a few years later. The foreign-interested genes that will be used in animal transgenic techniques are prepared using a variety of methods. The produced gene of interest is placed into a variety of vectors, including yeast artificial chromosomes, bacterial plasmids, and cosmids. Several techniques, including heat shock, electroporation, viruses, the gene gun, microinjection, and liposomes, are used to deliver the created vector, which includes the interesting gene, into the host cell. Transgenesis can be carried out in the gonads, sperm, fertilized eggs, and embryos through DNA microinjection, retroviruses, stem cells, and cloning. The most effective transgenic marker at the moment is fluorescent protein. Although transgenesis raises a number of ethical concerns, this review concentrates on the fundamentals of animal transgenesis and its usage in industry, medicine, and agriculture. Transgenesis success is confirmed by the integration of an antibiotic resistance gene, western and southern blots, PCR, and ELISA. If technology solves social and ethical problems, it will be the most promising in the future.

Background: MM (multiple myeloma) is a bone marrow disease with the accumulation of malignant plasma cells characterized by the neoplastic transformation of differentiated B cells. The onset and progression of cancer are greatly influenced by telomere dysfunction. We aimed to study the biomarker potential and prognostic significance of shelterin complex and hTERT. Telomere length and gene expression were measured using real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR), and these results were further correlated with clinical parameters.

Results: Our study showed increased expression of all genes in complex, hTERT, and TL in MM (n = 72) in comparison with controls (n = 31). TRF2 (P = 0.025) and hTERT (P = 0.0002) displayed significant association among cytogenetic analysis. The receiver operative curve showed POT1 and RAP1 with a greater area under the curve (AUC). RAP1 (P = 0.020) and hTERT (P = 0.037) displayed to be independent prognostic markers for overall survival. Clinical parameters and genes were observed to be significantly correlated.

Conclusion: Our study findings showed variation in telomere-associated genes and suggest the participation of these genes as prognostic markers in MM. These results all together highlight the evaluation and role of genes involved in telomeric alteration and TL, providing the opportunity to study new therapeutic approaches in patients with MM.

Background: Finding natural products with anticancer activity is an effective strategy to fight this disease. In this respect, Lepidium sativum or garden cress (family Brassicaceae) has been widely used worldwide for its wide therapeutic application, including anticancer and chemoprotective agents. Plant tissue culture techniques hold great promise for natural product enhancement without any climatic boundaries. In this study, glucosinolates and petroleum ether fractions were isolated from in vitro cell cultures and used against different carcinoma cell lines to investigate their anticancer potential.

Methods: In this study, callus cultures from leaf and root explants were initiated, cell suspension cultures were established, and cell growth and viability profiles were characterized. Different amino acids were added as precursors to the cell suspension cultures to enhance glucosinolates accumulation. Gas chromatography-mass spectrometric analysis (GC-MS) of glucosinolates and petroleum ether fractions was performed, and all fractions were tested against different carcinoma cell lines.

Results: The findings clarified that the maximum callus initiation percentage was obtained in the medium containing 1.0 mg/l 2,4-dichlorophenoxy acetic acid (2,4-D) + 1.0 mg/l kinetin (Kin) (C1). The viable cell number of cell suspension cultures from leaves and roots increased until it reached the maximum values on day 15. Adding tyrosine and methionine to the cell suspension cultures was the most influential and recorded high glucosinolate percentages. 1H-Cyclopenta (b) pyridine-3-carbonitrile-4,5,6,7-tetrahydro-2-methylthio-4-spirocyclohexane was the main glucosinolate compound found in tyrosine-treated leaf suspension (GLT). Fifteen compounds were detected in the petroleum ether fraction in both cell suspensions initiated from the leaf and root (OL and OR). The major compounds were benzene-1,3,5-trimethyl (12.99%) in root cell suspension (OR), and benzene-2-ethyl-1,4-dimethyl (10.66%) in leaf cell suspension (OL). All glucosinolate extracts demonstrated significant anticancer activity against the prostate (PC3), lung (A-549), colorectal (caco2), and liver (HepG2) cell lines. Glucosinolates extracted from leaf cell suspension (GL) were the most active on the hepatocellular carcinoma cell line (HepG2) among all remaining glucosinolate extracts. Treated hepatocellular carcinoma with an IC₅₀ of GL extract (47.5 ug/ml) upregulates pro-apoptotic BAX and downregulates anti-apoptotic BCL2, which disrupts the BAX/BCL2 ratio, leading to activation of caspase 3 inside treated HepG2 cells.

Conclusions: The anticancer action of the GL extract was validated by the cell cycle study of its glucosinolates, which successfully promoted apoptosis and reduced hepatocellular growth by causing S-phase arrest.

Background: Probiotics are live microorganisms that provide beneficial effects on the host's health when exploited in adequate amounts. This study aimed at carrying out whole-genome sequence analysis and in vitro potential probiotic characteristics of Lactococcus lactis subsp. lactis strain Lac3 isolated from the spontaneously fermented buffalo milk named Dadih.

Results: The results from de novo assembly indicated that the assembled genome consisted of 55 contigs with a genome size of 2,441,808 bp ~ (2.44 Mb), and GC % content of 34.85%. The evolution history result showed that the strain Lac3 was closely related to Lactococcus lactis species deposited in NCBI with a sequence similarity ≥ 99.93%. L. lactis subsp. lactis Lac3 was non-pathogenic with a probability of 0.21 out of 1 and had a pathogenicity score of zero (0), and neither harbored virulence factors nor acquired antibiotic resistance phenotypes. L. lactis subsp. lactis Lac3 exhibited the potential probiotic characteristics to tolerate acid at pH (2.0 and 5.0), salinity (1-5% NaCl), bile salt of (0.3-1.0%) and had auto-aggregation capacity increased from 6.0 to 13.1%.

Conclusion: This study described a novel strain of Lactococcus lactis subsp. lactis called Lac3, which exhibits probiotic properties that could be beneficial in the development of probiotics.

Background: Eighty-three strains of Leuconostoc mesenteroides were isolated from Algerian raw camel milk. Based on morphological, biochemical, and physiological characters tests, strains were identified as Ln. mesenteroides subsp. mesenteroides. Seven strains had a remarkable antagonistic and probiotic characterization. The present study aims at identifying these strains by means of 16 s rRNA gene sequencing and Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), extending phenotypic and genotypic studies done previously.

Results: The phyloproteomic dendrograms of the studied strains based on MALDI-TOF MS provided the same identification with more intraspecific information from the 16S rRNA gene sequencing based on phylogenetic analysis. The latter were in agreement with the previous biochemical/physiological identification, the seven isolated strains were Ln. mesenteroides subsp. mesenteroides.

Conclusions: Remarkably, MALDI-TOF MS fingerprinting was found to be effective enough as 16S rRNA gene sequencing identification, allowing faster and more reliable analysis than biochemical/physiological methods.

Background: Hydrolytic enzymes from halophilic microorganisms have a wide range of industrial applications. Herein, we report the isolation of Halobacillus sp. HAL1, a moderately halophilic bacterium that produces a novel high molecular weight extracellular alkaline protease when grown in fish processing wastes as a substrate.

Results: Results showed that the isolated strain belonged to the genus Halobacillus, and it was designated as Halobacillus sp. HAL1 with the GenBank accession number OK001470. The strain secreted an extracellular alkaline protease, and the highest yield was obtained when it was grown in a medium with fish wastes substrate as the sole nutritional source (10 g/L) and incubated at 25 °C under shaking conditions. The enzyme was partially purified by Sephadex G-100 column chromatography. Zymographic analysis showed two casein degrading bands of about 190 and 250 KDa. The optimum enzyme activity was at a temperature of 50 °C at pH 8. The proteolytic activity was enhanced in the presence of metal ions (Ca²⁺, Mg²⁺, and Mn²⁺), surfactants (Tween 80, SDS, and Triton-X100), H₂O₂, and EDTA.

Conclusion: Our study indicates that Haobacillus sp. HAL1 is a moderately halophilic strain and secrets a novel high molecular wight alkaline protease that is suitable for detergent formulation.

Background: Zinc oxide nanoparticles (ZnO NPs) can be considered as nanofertilizer providing zinc as an essential micronutrient for plant growth and production at specific safe dose, however, above this dose; ZnO NPs induce oxidative stress. The present research aimed to evaluate some physiological and molecular effects of ZnO NPs on Trigonella foenum-graecum (fenugreek) plant.

Results: The ZnO NPs were applied at five different concentrations (10, 20, 30, 40, and 50 mg/l) via soaking fenugreek seeds for 24 h. Fenugreek seedlings were harvested after 14 days for biomass and biochemical analyses. The results revealed that increasing ZnO NPs concentration led to a significant increase in all measured parameters until peaked at 30 mg/l; after that, a decline trend was detected. However, malondialdehyde (MDA) increased significantly just at higher concentrations of ZnO NPs (40 and 50 mg/l). In addition, genetic variation measure using start codon targeted (SCoT) markers revealed that ZnO NP treatments exhibited limited genetic variation.

Conclusion: Results showed that treatment with ZnO NPs at 30 mg/l can improve biomass, bioactive compounds, and antioxidant activity of fenugreek seedlings, besides being safe for DNA. So, this concentration could be a decent nanofertilizer for fenugreek plant.

Background: Prostate cancer (PC) is a silent but potent killer among men. In 2018, PC accounted for more than 350, 000 death cases while more than 1.2 million cases were diagnosed. Docetaxel, a chemotherapeutic drug belonging to the taxane family of drugs, is one of the most potent drugs in combating advanced PC. However, PC cells often evolve resistance against the regimen. Hence, necessitating the search for complementary and alternative therapies. Quercetin, a ubiquitous phytocompound with numerous pharmacological properties, has been reported to reverse docetaxel resistance (DR) in docetaxel-resistant prostate cancer (DRPC). Therefore, this study aimed to explore the mechanism via which quercetin reverses DR in DRPC using an integrative functional network and exploratory cancer genomic data analyses.

Results: The putative targets of quercetin were retrieved from relevant databases, while the differentially expressed genes (DEGs) in docetaxel-resistant prostate cancer (DRPC) were identified by analysing microarray data retrieved from the Gene Expression Omnibus (GEO) database. Subsequently, the protein-protein interaction (PPI) network of the overlapping genes between the DEGs and quercetin targets was retrieved from STRING, while the hub genes, which represent the key interacting genes of the network, were identified using the CytoHubba plug-in of Cytoscape. The hub genes were further subjected to a comprehensive analysis aimed at identifying their contribution to the immune microenvironment and overall survival (OS) of PC patients, while their alterations in PC patients were also revealed. The biological roles played by the hub genes in chemotherapeutic resistance include the positive regulation of developmental process, positive regulation of gene expression, negative regulation of cell death, and epithelial cell differentiation among others.

Conclusion: Further analysis revealed epidermal growth factor receptor (EGFR) as the most pertinent target of quercetin in reversing DR in DRPC, while molecular docking simulation revealed an effective interaction between quercetin and EGFR. Ultimately, this study provides a scientific rationale for the further exploration of quercetin as a combinational therapy with docetaxel.

Background: Monkeypox virus is a small, double-stranded DNA virus that causes a zoonotic disease called Monkeypox. The disease has spread from Central and West Africa to Europe and North America and created havoc in some countries all around the world. The complete genome of the Monkeypox virus Zaire-96-I-16 has been sequenced. The viral strain contains 191 protein-coding genes with 30 hypothetical proteins whose structure and function are still unknown. Hence, it is imperative to functionally and structurally annotate the hypothetical proteins to get a clear understanding of novel drug and vaccine targets. The purpose of the study was to characterize the 30 hypothetical proteins through the determination of physicochemical properties, subcellular characterization, function prediction, functional domain prediction, structure prediction, structure validation, structural analysis, and ligand binding sites using Bioinformatics tools.

Results: The structural and functional analysis of 30 hypothetical proteins was carried out in this research. Out of these, 3 hypothetical functions (Q8V547, Q8V4S4, Q8V4Q4) could be assigned a structure and function confidently. Q8V547 protein in Monkeypox virus Zaire-96-I-16 is predicted as an apoptosis regulator which promotes viral replication in the infected host cell. Q8V4S4 is predicted as a nuclease responsible for viral evasion in the host. The function of Q8V4Q4 is to prevent host NF-kappa-B activation in response to pro-inflammatory cytokines like TNF alpha or interleukin 1 beta.

Conclusions: Out of the 30 hypothetical proteins of Monkeypox virus Zaire-96-I-16, 3 were annotated using various bioinformatics tools. These proteins function as apoptosis regulators, nuclease, and inhibitors of NF-Kappa-B activator. The functional and structural annotation of the proteins can be used to perform a docking with potential leads to discover novel drugs and vaccines against the Monkeypox. In vivo research can be carried out to identify the complete potential of the annotated proteins.