The yidRv gene, isolated from the hypervirulent Klebsiella pneumoniae (hvKP), is a novel gene with an unknown function; however, it has exhibited high homology to the yidR, a gene recognized as potential vaccine candidate. The aim of this study was to clone the yidRv gene from the Indonesian hvKP and to investigate its overexpression in Escherichia coli. In the experiment, yidRv was cloned into pET21 to construct pYik23. Recombinant protein YidRv was produced by growing E. coli BL21 (DE3)/pYik23 in LB medium with ampicillin at 29 °C, inducing protein synthesis with 0.5 mM IPTG for 20 hours. Purification was performed using Ni‐NTA resin, and the purified protein (50 µg) was administered to BALB/c mice to test for the production of IgG, IgM and IgA on 2 days before and day 19th and 37th after the first vaccination. The results show a significant induction of IgG and IgM, but not of IgA antibodies. In conclusion, the yidRv gene was overexpressed in E. coli BL21 (DE3) at high levels in soluble form, with the recombinant protein able to be purified to 90% homogeneity. The recombinant YidRv demonstrated the ability to stimulate the generation of both IgM and IgG antibodies.

Recent advances in biotechnology have sparked global interest in developing biosimilar drugs, particularly those containing physiologically active proteins, such as growth factors and cytokines. The methylotrophic yeast Pichia pastoris can produce and secrete fully active heterologous proteins with strong secretory capacity and low levels of native proteins and has the ability to achieve high cell densities. In this study, a yeast‐based system was used to express and purify recombinant human granulocyte colony‐stimulating factor (rG‐CSF). Cultures were induced every 12 h for 48 h to express rG‐CSF, and parameters such as cell density, media pH, and cell dry weight were observed. Cell density increased along with the corresponding secretion of rG‐CSF during the induction period, as determined by Western blot assay, while the pH of the media remained stable. Ammonium sulfate at different saturation levels was used to precipitate the recombinant protein, with the highest total protein content determined spectrophotometrically at 29.6 µg/mL. Ni‐NTA resin with affinity column purification was used to purify the recombinant protein. The purified protein showed rG‐CSF with a molecular weight of approximately 18 kDa based on SDS‐PAGE analysis and immuno slot blot assay detected in purple. Overall, the study results indicated that the production and purification of rG‐CSF was successful, although optimization was required. The long‐term goal of this research is to discover alternative methods and sources for producing biosimilars of the therapeutic protein rG‐CSF, which can be utilized in the pharmaceutical industry to support health programs, particularly cancer treatment.

Given the current status of climate change and its impact on global food security, it is imperative to improve the abiotic stress tolerance of crop plants to enhance productivity. Traditional plant breeding methods have been widely employed to develop climate‐resilient crops; however, their success has been limited due to the lack of understanding of the complex relationships between genes and stress‐related phenotypes. The advent of modern genomics has enabled the expression analysis of stress genes in plants, as genome‐wide information is readily accessible and can be utilized to assign and validate the gene functions. This article highlights the potential applications and limitations of present‐day genomic technologies based on genome mapping, gain or loss‐of‐function analysis for identification of the role of a particular gene in abiotic stress response in plants. Such technologies are highly efficient in candidate gene identification; gene‐trait relationships establishment; functional elucidation of genes; and stress genes modification in crop plants. Modern high throughput genomic technologies offer wide scope for deciphering the complexities of genetic regulation of stress in plants; modulating stress responses; and developing stress tolerance in crop plants against drought, temperature, salinity, osmotic imbalance, herbicides and heavy metal toxicity.

Dendrobium ovatum is a tropical epiphytic orchid endemic to the Western Ghats of India and has been listed as a threatened species in recent research due to its declining populations and changes in flowering and fruit set patterns. This study aims to investigate the mycoflora associated with the roots, stems and leaves of D. ovatum. Both surface‐associated and endophytic fungal associates were isolated and identified using morphological and molecular methods. The study resulted in the isolation of 139 cultures, which were divided into 24 morphotypes, 99% of which belonged to Ascomycota. The most dominant members, Trichoderma harzianum and Colletotrichum gloeosporioides, were consistently observed across all the study sites. Tissue‐specific fungal diversity analysis revealed that each organ was dominated by a distinct fungal group, forming characteristic communities specific to each tissue. The roots of D. ovatum exhibited the highest species richness and diversity, compared to the stem and leaves. This research also represents the first documentation of fungal associates of the threatened orchid D. ovatum.

This study investigated the diversity of bacterial community in the samples of pigeon pea (Cajanus cajan L. Millsp.) soaked in water for 12 h and 24 h. The detection of certain bacterial species in the samples that can be isolated and potentially be used as starter cultures in the development of pigeon pea‐based functional foods is the importance of this study. For bacterial identification, the V1–V9 regions on the 16S ribosomal RNA gene were amplified using 27F and 1492R primers under specific polymerase chain reaction conditions. Genomic DNA (130 ng) was sequenced on the R9.4 flow cell by Oxford Nanopore Technologies using a GridION sequencer. Library preparations were conducted using a Native Barcoding Kit 24 V14 (SQK‐NBD114.24). Primary data were acquired using MinKNOW version 22.05.7. A total of 13 bacterial families and 89 genera were identified in the pigeon pea sample soaked for 12 h, and 26 families and 90 genera were identified in the pigeon pea soaked for 24 h. The values of five diversity indices showed that the sample soaked in water for 24 h had richer bacterial abundance and diversity than for 12 h. Shannon and Simpson values revealed the higher bacterial diversity in the samples collected at 24 h than in those collected at 12 h. Species observation and abundance‐based coverage estimators (ACE) values demonstrated that the samples collected at 24 h harbored higher bac‐ terial richness than those collected at 12 h. Bacterial communities during soaking of the pigeon pea were dominated by the family Enterobacteriaceae and genus Enterobacter. The presence of bacterial genera like Lacticaseibacillus, Lentilactobacillus, and Secundilactobacillus is interesting because of their importance as starter cultures for fermented plant‐based milk products

Transient gene expression is an approach used to study transient genes across various species, with infiltration by Agrobacterium tumefaciens (agroinfiltration) being a commonly used method. Agroinfiltration offers a simple and effective means of delivering transgenes into the plant genome. An alternative method for enhancing the quality and productivity of orchids as ornamental plants is genetic modification through agroinfiltration. Although Agrobacterium‐mediated genetic transformation by immersion has been used on the Phalaenopsis amabilis (L.) Blume species of orchid, transformation efficiency using the immersion technique remains relatively low and the method itself is challenging due to its requirement for aseptic handling. The application of agroinfiltration in P. amabilis has not previously been reported. This study investigates the impact of the injection site, acetosyringone concentration, bacterial density (OD600), and injection volume to determine the optimum conditions for agroinfiltration on P. amabilis. The results demonstrated that injection site had a noticeably distinct impact on transformation effectiveness, with the abaxial position of the leaf being the optimal site for Agrobacterium culture suspension injection. While adjustments in acetosyringone concentration, bacterial density (OD600), and injection volume did not significantly affect transformation efficiency, they did influence the peak time of GFP fluorescence. Acetosyringone at a concentration of 200 µM, an OD600 of 1.0 for Agrobacterium culture, and an injection volume of 500 µL effectively accelerated GFP expression duration.

The PhoR‐PhoP two‐component regulatory system, which is responsible for regulating the virulence of Mycobacterium tuberculosis, presents a promising target for the development of novel tuberculosis drugs. Disrupting the interaction of PhoR‐PhoP proteins has the potential to decrease the virulence of the bacterium, rendering it more vulnerable to immune system clearance. A dimer‐based screening system was developed to screen for inhibitors of PhoR dimerization. The coding sequence for the cytoplasmic domain of PhoR (cytoPhoR) was combined with the DNA‐binding domain of the AraC repressor coding sequence. These sequences were positioned upstream of the emerald green fluorescent protein (EmGFP), which serves as a reporter gene. and controlled by the araC promoter. The in silico investigation examined the modeling of the fusion AraC_cytoPhoR and its binding to the promoter. The plasmid construct generated, namely pAraC_PhoRMTB, was synthesized and confirmed using DNA sequencing. The confirmed plasmid was then transformed into Escherichia coli BL21(DE3). Both SDS PAGE and fluorescence analysis indicated that the transformed culture expressed the AraC‐cytoPhoR fusion protein and displayed lower relative fluorescence in comparison to the transformed culture consisting solely of the AraC DNA‐binding domain coding sequence. This reduction in fluorescence suggests that the dimer‐based screening system effectively monitors the inhibition of dimerization of cytoPhoR. These analysis findings indicate that the system is now ready for use in the screening of PhoR dimerization inhibitors.

Prolonged exposure to high‐intensity UVB induces the formation of reactive oxygen species (ROS) in skin tissue, triggering an increase in matrix metalloproteinase‐3 (MMP‐3) enzyme production and leading to collagen degradation. Moringa oleifera (MO) contains bioactive compounds known for ROS‐scavenging and anti‐inflammatory properties. However, the precise molecular mechanism of action remains unclear, requiring the inhibition of MMP‐3 activation and regulation of collagen deposition. This study aims to elucidate the potential effect of MO leaf extract‐based gel in restoring collagen deposition by reducing MMP‐3 activation in UVB irradiate‐induced collagen loss in rats. This study employed a completely randomized design, comprising four groups: a healthy group without UVB radiation, a negative control group subjected to UVB radiation and receiving a placebo, and two treatment groups exposed to UVB radiation with 5% or 10% moringa leaf extract‐based gel (MO‐5% or MO‐10%), respectively. Results showed that MO‐5% and MO‐10% significantly reduced MMP‐3 gene expression and increased collagen density compared to the negative control group (p < 0.05). Moringa oleifera leaf extract ameliorates collagen degradation by inhibiting MMP‐3 expression in UVB‐induced rats, suggesting its potential as a pharmacological and cosmetic agent for UVB‐induced skin damage.

In addition to the issue of pork contamination, processed meats frequently contain traces of rat meat. Therefore, detection and quantification of the pork and rat DNA in cases of meat and processed meat adulteration are necessary. In the current study, two gene targets of the cytochrome b for pigs and the Mt‐atp6 of Rattus norvegicus for rats were used in the absolute multiplex quantitative real‐time PCR (m‐qPCR). The sample DNA was amplified with a standard as positive control in the various concentration of 1000 pg, 100 pg, 10 pg, 0.1 pg, 0.01 pg, and 0.001 pg. There were 25 processed meat samples and 5 fresh meat samples identified in this study. Among the total of 30 samples assessed, 6 samples were successfully detected and quantified their pork and rat DNA contamination. One sample was contaminated with pork DNA with a concentration of 2.451×10‐4 pg (“Meatball 3). Five samples were contaminated with rat DNA with a concentration of 3.603×10‐11 pg (“Sempol 3”), 2.196×10‐10pg (“Meatball 6”), 4.908×10‐11 pg (“Siomay 3”), 1.489×10‐10 pg (“Grinding 2”), and 3.564×10‐10 pg (“Grinding 4”). In this study, we have discovered that the contamination of pork and rat were detected in the samples. It suggested that this method is applicable for detecting the contaminant in processed meat samples

The determination of transcript accumulation values significantly affects gene expression in oil palm. Various genes are involved in pathogen infection, including probable 2‐oxoglutarate‐dependent dioxygenase At5g05600 (EgUnk3), zinc finger protein 2‐like (EgZFP2), and inositol polyphosphate multikinase beta‐like (EgIPK2b). Gene expression is typically measured using relative quantitative methods to calculate differences in quantitative values in the expression levels of targeted genes compared to a reference gene. However, the effectiveness of these methods in assessing the expression of EgUnk3, EgZFP2, and EgIPK2b, which are involved in Ganoderma boninense infection in oil palm seedlings, requires evaluation. This study aimed to establish an effective and straightforward method for analyzing the expression of EgUnk1, EgZFP2, and EgIPK2b genes in oil palm seedlings infected with G. boninense, utilizing Ct value correction through regression coefficients on the 2‐ΔΔCt and E‐ΔΔCt approaches. A correlation regression revealed values of 0.28, ‐0.32, and 0.29 for delta Ct of EgUnk1, EgZFP2, and EgIPK2b, respectively. However, a negative correlation in the Ct mean was corrected by linear regression for the targeted genes: ‐0.55, ‐0.81, and ‐0.29 for EgUnk1, EgZFP2, and EgIPK2b, respectively. The amplification factor (E) and efficiency value (R) using the EgActin gene were 1.95 and 94.92%, respectively. Normalization of log10 on the fold change value 2‐ΔΔCt and 1.95‐ΔΔCt approaches using the regression coefficient yielded consistent results for the EgUnk1, EgZFP2, and EgIPK2b genes. Overall, EgUnk3 and EgIPK2b genes exhibited downregulated expression in susceptible oil palm seedlings (‐0.60 for 2(‐ΔΔCt) and ‐0.58 for 1.95(‐ΔΔCt)), whereas EgIPK2b gene showed up‐regulated and the highest value in inoculated resistant seedlings (1.39 for 2(‐ΔΔCt) and 1.34 for 1.95(‐ΔΔCt)). Basal stem rot disease (BSR) in oil palm decreased EgUnk1 and EgIPK2b expression in susceptible seedlings but increased EgZFP2 gene expression in resistant ones. The results of this research provide valuable corrections to Ct values obtained directly from RT‐qPCR machines using simple linear regression. Consequently, the Ct values of target genes and reference genes exhibit smaller bias values, rendering gene expression levels more reliable.