Conservation and function of Dazl in promoting the meiosis of goat male germline stem cells

Dazl (deleted in azoospermia-like) is a conserved gene in mammalian meiosis, which encodes RNA binding protein required for spermatocyte meiosis. Up to date, the expression and function of Dazl in the goat testis are unknown. The objectives of this study were to investigate the expression pattern of Dazl in dairy goat testis and their function in male germline stem cells (mGSCs). The results first revealed that the expression level of Dazl in adult testes was significantly higher than younger and immature goats, and azoospermia and male intersex testis. The dairy goat Dazl is highly conserved analysed by several online and bioinformatics software, respectively. Over-expression of Dazl promoted the expression of meiosis-related genes in dairy goat mGSCs. The expression of Stra8 was up-regulated by over-expression of Dazl analysed by Luciferase reporter assay. Taken together, results suggest the Dazl plays an important role in dairy goat spermatogenesis and that over-expression of Dazl may promote Stra8 expression in dairy goat mGSCs.     

Simultaneous enhanced catalytic activity and thermostability of a 1,3-1,4-β-glucanase from Bacillus amyloliqueformis by chemical modification of lysine residues

The thermostablility and enzymatic activity of 1,3-1,4-β-glucanase (BglA) from Bacillus amyloliquefaciens was improved by modifying five (out of 12) ε-amino groups in lysine residues with nitrous acid. The optimal modification condition for BglA was determined as 30 mM nitrous acid at, 40 °C for 30 min. The optimally-modified BglA had higher specific activity and T ₅₀ value, which were 3,370 U/mg and 70 °C, respectively. Its half-life values at 50 and 60 °C were extended and reached 58.5 and 49.5 min, respectively. Circular dichroism analysis showed that the secondary structures in modified BglA were almost the same with that of wild-type BglA. Thus, modification of lysine residues can simultaneously improve the activity and thermostability of β-glucanase which are ideal targets for further protein engineering.     

Field evaluation of attractive lures for the fruit fly Bactrocera minax (Diptera: Tephritidae) and their potential use in spot sprays in Hubei Province (China)

The Chinese citrus fruit fly, Bactrocera minax (Enderlein) is a univoltine Tephritidae pest that infests Citrus species. Field trials were conducted in 2010 to determine the potential use of a lure based on enzymatical-hydrolyzed beer yeast as liquid bait (hereafter named H-protein bait) for B. minax in the Hubei province, China. In a citrus orchard, we compared the attractiveness among aqueous solutions of H-protein bait, GF-120 fruit fly bait, sugar-vinegar-wine mixture, torula yeast, and Jufeng attractant when used in traps and in spot sprays, that is, lures used in combination with the insecticide trichlorphon. The H-protein bait was the most attractive lure in traps, ensnaring significantly more adults than sugar-vinegar-wine mixture, torula yeast, and Jufeng attractant, in decreasing efficiency order. In spot sprays those with H-protein bait killed significantly more female and male flies within 40 min than those with sugar-vinegar-wine mixture, GF-120, Jufeng attractant, and the control. In addition, the total number of flies killed by H-protein bait during the spot spray duration was higher than other treatments. Our results demonstrated that the H-protein bait may be a useful tool in citrus orchards in China to monitor B. minax populations as well as to manage this pest when used in spot sprays.     

Gut microbiota in wild and captive Guizhou snub‐nosed monkeys,Rhinopithecus brelichi

Many colobine species—including the endangered Guizhou snub‐nosed monkey (Rhinopithecus brelichi) are difficult to maintain in captivity and frequently exhibit gastrointestinal (GI) problems. GI problems are commonly linked to alterations in the gut microbiota, which lead us to examine the gut microbial communities of wild and captive R. brelichi. We used high‐throughput sequencing of the 16S rRNA gene to compare the gut microbiota of wild (N = 7) and captive (N = 8) R. brelichi. Wild monkeys exhibited increased gut microbial diversity based on the Chao1 but not Shannon diversity metric and greater relative abundances of bacteria in the Lachnospiraceae and Ruminococcaceae families. Microbes in these families digest complex plant materials and produce butyrate, a short chain fatty acid critical to colonocyte health. Captive monkeys had greater relative abundances of Prevotella and Bacteroides species, which degrade simple sugars and carbohydrates, like those present in fruits and cornmeal, two staples of the captive R. brelichi diet. Captive monkeys also had a greater abundance of Akkermansia species, a microbe that can thrive in the face of host malnutrition. Taken together, these findings suggest that poor health in captive R. brelichi may be linked to diet and an altered gut microbiota.     

SNPicker: a graphical tool for primer picking in designing mutagenic endonuclease restriction assays

Simple, low-cost and accurate genotyping methods for single nucleotide polymorphisms (SNPs) are in high demand in the post-genome-sequencing era. We present a graphical tool called SNPicker, implemented in Java, which significantly facilitates the design of mutagenic endonuclease restriction assays. SNPicker uses the online NEB REBASE to automatically scan for all possible designs of mutagenic primers that can facilitate the picking of mismatched PCR primers to artificially introduce or abolish a restriction site at the target SNP site. We successfully applied SNPicker in designing endonuclease restriction assays for 14 SNPs for the MTHFR gene, the Coagulation Factor II gene and the Coagulation Factor V gene. The SNP assays designed using SNPicker were cross-validated using the MassARRAY technology.     

Colorimetric method for identifying plant essential oil components that affect biofilm formation and structure

The specific biofilm formation (SBF) assay, a technique based on crystal violet staining, was developed to locate plant essential oils and their components that affect biofilm formation. SBF analysis determined that cinnamon, cassia, and citronella oils differentially affected growth-normalized biofilm formation by Escherichia coli. Examination of the corresponding essential oil principal components by the SBF assay revealed that cinnamaldehyde decreased biofilm formation compared to biofilms grown in Luria-Bertani broth, eugenol did not result in a change, and citronellol increased the SBF. To evaluate these results, two microscopy-based assays were employed. First, confocal laser scanning microscopy (CLSM) was used to examine E. coli biofilms cultivated in flow cells, which were quantitatively analyzed by COMSTAT, an image analysis program. The overall trend for five parameters that characterize biofilm development corroborated the findings of the SBF assay. Second, the results of an assay measuring growth-normalized adhesion by direct microscopy concurred with the results of the SBF assay and CLSM imaging. Viability staining indicated that there was reduced toxicity of the essential oil components to cells in biofilms compared to the toxicity to planktonic cells but revealed morphological damage to E. coli after cinnamaldehyde exposure. Cinnamaldehyde also inhibited the swimming motility of E. coli. SBF analysis of three Pseudomonas species exposed to cinnamaldehyde, eugenol, or citronellol revealed diverse responses. The SBF assay could be useful as an initial step for finding plant essential oils and their components that affect biofilm formation and structure.     

Three Meta-Analyses Define a Set of Commonly Overexpressed Genes from Microarray Datasets on Astrocytomas

Glioma is one of the most common tumors of the central nervous system, and one of its main types is astrocytoma. Microarray technology has been widely used to explore the molecular mechanism of cancer. It is universally accepted that meta-analysis considerably improves the statistical robustness of results, particularly in clinical research. To obtain the maximum reliability, we used three different meta-analyses to integrate the four microarray datasets, GSE16011, GSE4290, GSE2223, and GSE19728 (local), and defined the common differentially expressed genes (DEGs) in astrocytomas compared with normal brain tissue. Four DEGs, PCNA, CDC2, CDK2 and CCNB2, which are components of the cell cycle pathway, were chosen for Real-Time Polymerase Chain Reaction (RT-PCR) and immunohistochemistry validation. PCNA is similar to the P53 gene and has been widely implicated in various cancers including gliomas. Therefore, the expression status of PCNA in our study was considered as a reference to test our whole experimental scheme, and the results indicate that our methodology is valid. Although a few studies have reported the overexpression of the CDC2, CDK2 and CCNB2 genes in glioma cell lines, we are the first to identify the statuses of these genes in human astrocytoma tissues at the mRNA and protein levels. The results of the gene validations strongly suggested that the genes play an important role in astrocytomas and could potentially be valuable in the diagnosis and treatment of astrocytoma.