The Use of Zinc Finger Protein Gene to Detect the Presence of Y Chromosome in the Cow

Total genomic DNA from 24 normal cows and a normal male cattle were isolated from leucocyte preparation. Polymerase Chain Reaction (PCR) amplification of these genomic DNA using the PI-5EZ and P2-3EZ primers for ZFX/ZFY gene resulted in a fragment of approximately 440 bp in size from both male and female samples. This PCR product represents part of the ZFX/ZFY gene defined by the primers used. The PCR products were proven to be amplified from bovine genomic DNA from the dot blot hybridization studies. After Pst I digestion, the female samples (ZFX) remained undigested while the male (ZFX/ZFY) showed three fragments of approximately 440 bp, 340 bp and 100 bp. Thus the ZFY gene has a unique Pst I site. The PCR product from a population of 45 female and 47 male cattle was further tested for the presence of the Pst I cleavage. There was no Pst I site in all the female (ZFX) samples, while 45 out of 47 samples from the male were cleaved. From X2 analysis of the data, the Pst I cleavage on ZFY was significantly sex dependent. The restriction analysis of ZFX/ZFY gene showed the existence of a unique Mnl I site on ZFX and more than one Alu I restriction site on both ZFX and ZFY. The PCR product from an infertile female cattle (supposedly, 60, XX and therefore ZFX/ZFX) was subjected to the Pst I digestion, and found to be cleaved indicating the presence of the ZFY gene and possibly the Y chromosome. The latter was confirmed by cytogenetic analysis. Post mortem and autopsy evaluation of the animal showed a partial development of male characteristic and a defective female reproduction system The use of the peR diagnosis for the presence of the Y chromosome was applied to a population of 30 suspected infertile female animals, and two were found to have the ZFY gene indicating the possible presence of the Y chromosome. Thus, peR amplification of the ZFX/ZFY gene followed by Pst I digestion and analysis on gel electrophoresis can be used as an approach to establish the presence of the Y chromosome in an infertile female cattle. This approach however, needs to be supported by karyo typing to establish unequivocally the screening procedure

Antiproliferative effect of methanolic extraction of tualang honey on human keloid fibroblasts

<p>Abstract</p> <p>Background</p> <p>Keloid is a type of scar which extends beyond the boundaries of the original wound. It can spread to the surrounding skin by invasion. The use of Tualang honey is a possible approach for keloid treatment. The objective of this study was to determine the antiproliferative effect of methanolic extraction of Tualang honey to primary human keloid fibroblasts and to identify the volatile compounds in methanol extraction of Tualang honey.</p> <p>Methods</p> <p>Crude Tualang honey was extracted with methanol and then dried using rota vapor to remove remaining methanol from honey. Normal and keloid fibroblasts were verified and treated with the extracted honey. Cell proliferation was tested with [3-(4,5-dimethylthiazol-2-yi)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt] (MTS) assay. Extraction of Tualang honey using methanol was carried out and the extracted samples were analysed using gas chromatography-mass spectrometry (GC-MS). The result was analysed using SPSS and tested with Kruskal-Wallis and Mann-Whitney tests.</p> <p>Results</p> <p>Methanolic extraction of honey has positive anti proliferative effect on keloid fibroblasts in a dose-dependent manner. The presence of fatty acids such as palmitic acid, stearic acid, oleic acid, linoleic acid and octadecanoic acid may contribute to the anti-proliferative effect in keloid fibroblasts.</p> <p>Conclusions</p> <p>The methanolic honey extraction has an antiproliferative effect on keloid fibroblasts and a range of volatile compounds has been identified from Tualang honey. The antiproliferative effect of keloid fibroblasts towards Tualang honey may involve cell signaling pathway. Identifying other volatile compounds from different organic solvents should be carried out in future.</p

Overview of the main methods used to combine proteins with nanosystems: absorption, bioconjugation, and encapsulation

The latest development of protein engineering allows the production of proteins having desired properties and large potential markets, but the clinical advances of therapeutical proteins are still limited by their fragility. Nanotechnology could provide optimal vectors able to protect from degradation therapeutical biomolecules such as proteins, enzymes or specific polypeptides. On the other hand, some proteins can be also used as active ligands to help nanoparticles loaded with chemotherapeutic or other drugs to reach particular sites in the body. The aim of this review is to provide an overall picture of the general aspects of the most successful approaches used to combine proteins with nanosystems. This combination is mainly achieved by absorption, bioconjugation and encapsulation. Interactions of nanoparticles with biomolecules and caveats related to protein denaturation are also pointed out. A clear understanding of nanoparticle-protein interactions could make possible the design of precise and versatile hybrid nanosystems. This could further allow control of their pharmacokinetics as well as activity, and safety

YB-1 And CTCF Differentially Regulate The 5-HTT Polymorphic Intron 2 Enhancer Which Predisposes To A Variety Of Neurological Disorders.

Mutation or inappropriate expression of the serotonin transporter (5-HTT) gene has been postulated as a possible predisposing factor in many CNS-related disorders, including numerous studies of affective disorders. The human gene encompasses 15 exons spanning -31 kb on chromosome 17qll (Lesch et aI., 1994)

N,N′-(Ethane-1,2-di­yl)bis­(4-chloro­benzene­sulfonamide)

The title mol­ecule, C14H14Cl2N2O4S2, lies on an inversion center. The mol­ecule is twisted in the region of the sulfonamide group with a C—S—N—C torsion angle of −67.49 (16)°. In the crystal, mol­ecules are connected via inter­molecular N—H⋯O and weak C—H⋯O hydrogen bonds, forming layers parallel to (100)

Construction of multi-epitopes vaccine candidate against SARS-CoV-2 D614G variant

COVID-19 caused by the SARS-CoV-2 virus has become a real threat due to the emergence of new variants which are more deadly with higher infectivity. Vaccine constructs that target specific SARS-CoV-2 variants are needed for stemming COVID-19 fatality. The spike (S) glycoprotein is the major antigenic component that triggers the host immune response. Reverse vaccinology strategy was applied to the S protein of COVID-19 variant D614G to identify highly ranked antigenic proteins. In this study, a multi-epitope synthetic gene was designed using computational strategies for the COVID-19 D614G variant. The SARS-CoV-2 D614G variant protein sequence was retrieved from the NCBI database. The prediction of linear B-cell epitopes was carried out using Artificial Neural Network (ANN)-based ABCpred and BepiPred 2.0 software. The top 15 highly antigenic epitopes sequences were then selected. Propred 1 and Propred servers were used to identify major histocompatibility complex (MHC) class I and class II binding epitopes within pre-determined B-cell epitopes to predict T-cell epitopes. The top 5 MHC class I and class II were selected. Further in-silico testing for its solubility, allergenicity, antigenicity, and other physiochemical properties was analyzed using Bpred. The constructed gene was subjected to assembly PCR and the gene product was confirmed by Sanger sequencing. The findings from this study suggested that a highly antigenic specific region of the SARS-CoV-2 D614G variant can be predicted in-silico and amplified using the assembly PCR method. The designed synthetic gene was shown to elicit specific humoral and cell-mediated immune responses towards the SARS-CoV-2 variants

Cytotoxicity Enhancement in MCF-7 Breast Cancer Cells with Depolymerized Chitosan Delivery of α-Mangostin

The application of α-mangostin (AMG) in breast cancer research has wide intentions. Chitosan-based nanoparticles (CSNPs) have attractive prospects for developing anticancer drugs, especially in their high flexibility for modification to enhance their anticancer action. This research aimed to study the impact of depolymerized chitosan (CS) on the cytotoxicity enhancement of AMG in MCF-7 breast cancer cells. CSNPs effectivity depends on size, shape, crystallinity degree, and charge surface. Modifying CS molecular weight (MW) is expected to influence CSNPs’ characteristics, impacting size, shape, crystallinity degree, and charge surface. CSNPs are developed using the method of ionic gelation with sodium tripolyphosphate (TPP) as a crosslinker and spray pyrolysis procedure. Nanoparticles’ (NPs) sizes vary from 205.3 ± 81 nm to 450.9 ± 235 nm, ZP charges range from +10.56 mV to +51.56 mV, and entrapment efficiency from 85.35% to 90.45%. The morphology of NPs are all the same spherical forms. In vitro release studies confirmed that AMG–Chitosan–High Molecular Weight (AMG–CS–HMW) and AMG–Chitosan–Low Molecular Weight (AMG–CS–LMW) had a sustained-release system profile. MW has a great influence on surface, drug release, and cytotoxicity enhancement of AMG in CSNPs to MCF-7 cancer cells. The preparations AMG–CS–HMW and AMG–CS–LMW NPs considerably enhanced the cytotoxicity of MCF-7 cells with IC50 values of 5.90 ± 0.08 µg/mL and 4.90 ± 0.16 µg/mL, respectively, as compared with the non-nano particle formulation with an IC50 of 8.47 ± 0.29 µg/mL. These findings suggest that CSNPs can enhance the physicochemical characteristics and cytotoxicity of AMG in breast cancer treatment

Cytotoxicity Enhancement of α-Mangostin with Folate-Conjugated Chitosan Nanoparticles in MCF-7 Breast Cancer Cells

α-mangostin (AM) is a promising natural anticancer agent that can be used in cancer research. However, its effectiveness can be limited by poor solubility and bioavailability. To address this issue, chitosan-based nanoparticles (CSNPs) have been investigated as a potential delivery system to enhance the cytotoxicity to cancer cells and improve selectivity against normal cells. In this study, we developed folate-conjugated chitosan nanoparticles (F-CS-NPs) using a carbodiimide-based conjugation method to attach folate to chitosan (CS), which have different molecular weights. The NPs were crosslinked using tripolyphosphate (TPP) via ionic gelation. To characterize the F-CS-NPs, we utilized various analytical techniques, including transmission electron microscopy (TEM) to evaluate the particle size and morphology, Fourier-transform infrared spectroscopy (FTIR) to confirm the presence of functional groups, and ultraviolet-visible spectroscopy (UV-Vis) to measure the absorption spectrum and confirm the presence of folate. The particle size of AM-F-CS-NPs ranged from 180 nm to 250 nm, with many having favorable charges ranging from +40.33 ± 3.4 to 10.69 ± 1.3 mV. All NPs exhibited the same spherical morphology. The use of F-CS-NPs increased drug release, followed by a sustained release pattern. We evaluated the cytotoxicity of AM, AM-F-CS-HMW, and AM-F-CS-LMW NPs against MCF-7 cells and found IC50 values of 8.47 ± 0.49, 5.3 ± 0.01, and 4.70 ± 0.11 µg/mL, respectively. These results confirm the improved cytotoxicity of AM in MCF-7 cells when delivered via F-CS-NPs. Overall, our in vitro study demonstrated that the properties of F-CS-NPs greatly influence the cytotoxicity of AM in MCF-7 breast cancer cells (significantly different (p < 0.05)). The use of F-CS-NPs as a drug-delivery system for AM may have the potential to develop novel therapies for breast cancer