New antifungal flavonoid glycoside from Vitex negundo

Flavonoids are ubiquitous in photosynthesizing cells and are common part of human diet. For centuries, preparations containing these compounds as the principal physiologically active constituents have been used to treat human diseases. Increasingly, this class of natural products is becoming the subject of anti-infective research. Our bioactivity guided fractionation of ethanolic extract of leaves of Vitex negundo resulted in the isolation of new flavone glycoside (4) along with five known compounds 1-3, 5 and 6. All the isolated compounds were evaluated for their antimicrobial activities. The new flavone glycoside 4 and compound 5 were found to have significant antifungal activity against Trichophyton mentagrophytes and Cryptococcus neoformans at MIC 6.25 microg/ml.     

Boundaries of the Origin of Replication: Creation of a pET-28a-Derived Vector with p15A Copy Control Allowing Compatible Coexistence with pET Vectors

During our studies involving protein-DNA interactions, we constructed plasmid pSAM to fulfill two requirements: 1) to facilitate transfer of cloned sequences from widely used expression vector pET-28a(+), and 2) to provide a vector compatible with pBR322-derived plasmids for use in cells harboring two different plasmids. Vector pSAM is a pET-28a(+)-derived plasmid with the p15A origin of replication (ori); pET-28a(+) contains the pBR322 replicon that is incompatible with other pBR322-derived plasmids. By replacing the original pET-28a(+) replicon–comprising the ori, RNAI, RNAII, and Rom–with the p15A replicon, we generated pSAM, which contains the pET-28a(+) multiple cloning site and is now compatible with pBR322-derived vectors. Plasmid copy number was assessed using quantitative PCR: pSAM copy number was maintained at 18±4 copies per cell, consistent with that of other p15A-type vectors. Compatibility with pBR322-derived vectors was tested with pGEX-6p-1 and pSAM, which maintained their copy numbers of 49±10 and 14±4, respectively, when both were present within the same cell. Swapping of the ori is a common practice; however, it is vital that all regions of the original replicon be removed. Additional vector pSAMRNAI illustrated that incompatibility remains when portions of the replicon, such as RNAI and/or Rom, are retained; pSAMRNAI, which contains the intact RNAI but not ROM, lowered the copy number of pGEX-6p-1 to 18±2 in doubly transformed cells due to retention of the pET-28a(+)-derived RNAI. Thus, pSAMRNAI is incompatible with vectors controlled by the pBR322 replicon and further demonstrates the need to remove all portions of the original replicon and to quantitatively assess copy number, both individually and in combination, to ensure vector compatibility. To our knowledge, this is the first instance where the nascent vector has been quantitatively assessed for both plasmid copy number and compatibility. New vector pSAM provides ease of transferring sequences from commonly used pET-28a(+) into a vector compatible with the pBR322 family of plasmids. This essential need is currently not filled.     

Effect of sub-diaphragmatic vagotomy on gastric mucosal mast cell population in pylorus ligated rats.

Gastric mucosal mast cell population was studied following sub-diaphragmatic vagotomy in albino rats, 6 and 12 h after pylorus ligation. Sub-diaphragmatic vagotomy significantly increased the gastric mucosal mast cell population in both 6 and 12 h groups, the increase being more in the latter. The results suggest that the vagal impulses act on the gastric mucosal mast cells causing their degranulation. Following vagotomy the contents stay bound within the mast cells. Increase in mast cell population with the longer experimental situation was possibly due to the continuation of normal turnover of the mast cells in the gastric mucosa. The present study, however, does not lead to a conclusion that the vagal influence on mast cell population is similar throughout the gastro-intestinal tract.