A new heterobifunctional reagent for immobilization of biomolecules on glass surface

Synthesis of a new heterobifunctional reagent, [N-(2-trifluoroethanesulfonatoethyl)-N-(methyl)-triethoxysilylpropyl-3-amine] (NTMTA) is described for the immobilization of a variety of biomolecules on glass surface. Its triethoxysilyl group reacts with glass surface and trifluoroethanesulfonate ester structure reacts selectively with aminoalkyl/mercaptoalkyl function in biomolecules. The immobilization can be achieved by two ways involving two steps. The first route involves the reaction of NTMTA with glass beads followed by attachment of aminoalkyl- or mercaptoalkylated biomolecules. The second one involves the reaction of biomolecules, viz., oligonucleotides, proteins, etc., with NTMTA via their aminoalkyl or mercaptoalkyl functions to form a biomolecule conjugate, which is then reacted with glass beads (unmodified) to complete immobilization process. This has been demonstrated by successful immobilization of 5'-mercaptoalkyl- or aminoalkylated oligonucleotides and some commonly used enzymes on glass beads using NTMTA reagent.     

Efficient 1–5 regioselective acylation of primary hydroxyl groups of fermentative derived xylitol catalyzed by an immobilized Pseudomonas aeruginosa lipase

The experiment described in this paper synthesizes xylitol acylated products from fermentative derived xylitol and acid anhydrides of various chain lengths in the presence of Tetrahydrofuran (THF) and acetonitrile using immobilized Pseudomonas aeruginosa (PL) lipase as a biocatalyst (97% residual activity up to five cycles) at 37°C, 200 rpm. This study examines a number of different acid anhydrides for their highly selective and efficient lipase-catalyzed acylation of primary hydroxyl groups in xylitol. Of those studied, the best results are obtained with butanoic anhydride, 80.12% after 4 h in acetonitrile followed by vinyl acetate, which results in 77.79% conversion after 8 h of incubation in THF as analyzed through high performance liquid chromatography (HPLC).     

Analysis of short-term variation and long-term drift during reagent kit lot change in an NABL accredited clinical biochemistry laboratory

BackgroundKit lot change in clinical biochemistry labs leads to variations in patient results. This study planned to identify variations during 60 reagent lot changes in our laboratory during the period from June 2018 to May 2019.MethodsA statistical analysis was performed to identify the difference between patient samples results variations and QC results. The long term drift was analyzed using a regression test.ResultsThere was a significant difference between the patient and QC results in 16.7% of reagent lot changes. Moreover, the extent of variation in QC results was 3.3%. No long-term drift was seen in three analytes which were studied using regression analysis.ConclusionsOur results showed that, during reagent kit lot change, along with QC material, the patient samples should also be run in order to identify the variation. However, this practice is presently ignored by most of the laboratories. There was no accumulated effect in our laboratory due to reagent kit lot change.     

Genetic transformation of chickpea (Cicer arietinum L.) with insecticidal crystal protein gene using particle gun bombardment

Here, we report the establishment of an efficient particle gun bombardment mediated genetic transformation in chickpea (Cicer arietinum L.) using cryIAc gene of Bacillus thuringiensis. Explants were bombarded with recombinant plasmids engineered for the expression of cryIAc transgene in plants and stable transformants regenerated in presence of benzyladenine, kinetin and kanamycin. Transformation frequency showed dependence on explant type, cultivars, plasmids, helium pressure and microcarrier type used. Integration of transgenes was demonstrated using polymerase chain reaction and Southern blot hybridization approaches in T ₀ plants. The expression of CryIA(c) delta-endotoxin and GUS enzyme was ascertained by enzyme linked immunosorbent assay and histochemical assays, respectively. These transgenic plants (T ₀) showed more protection and high mortality for Heliothis armigera and Spodoptera litura larvae as compared to control plants. The results of the present study indicate that highest transformation frequency (18%) could be achieved by use of gold as a microcarrier in combination with helium pressure of 900 psi. Among the other factors tested, plasmid pHS 102 was the most efficient plasmid, while epicotyl explant was the best explant source for particle gun bombardment. Among the different cultivars of chickpea tested, cultivar ICCC37 and PG-12 produced higher frequency of transformation frequency compared to others.     

Maternal serum leptin during pregnancy and infant birth weight: The influence of maternal overweight and obesity

Objective:

Few studies have examined whether the distinct metabolic patterns found in obese and nonobese pregnant women have different effects on the growing fetus. Our objective was to estimate the influence of longitudinal variation in maternal serum leptin levels on variation in infant birth weight in overweight/obese versus normal‐weight women.

Design and Methods:

In a prospective cohort of 286 gravidas, maternal weight and serum leptin levels at 6–10, 10–14, 16–20, 22–26, and 32–36 weeks gestation were measured. Effects of leptin levels on infant birth weight adjusted for gestational age at delivery (aBW) were analyzed using a linear regression model that accounted for the relationship of time‐varying predictors to the log‐transformed leptin concentrations.

Results:

Different relationships of aBW to maternal serum leptin and its rate of change across pregnancy were exhibited by overweight/obese and normal‐weight gravidas. For normal‐weight women, aBW is not associated with either the magnitude of the logarithm of the leptin concentration or with its rate of change in either the first or second half of pregnancy. Conversely, for overweight/obese women, an increase in the rate of change in maternal serum leptin in the second half of pregnancy is significantly associated with a decrease in aBW. This effect is distinct from that of maternal weight.

Conclusion:

Differences in the effect of maternal serum leptin on fetal growth between overweight/ obese and normal‐weight women suggest metabolic and physiologic heterogeneity between these groups. Such differences may be involved in the long‐term physiologic effects of the obese intrauterine environment on the health of the offspring.     

Identification of bacteriophage K20 binding regions of OmpF and lipopolysaccharide in Escherichia coli K-12

Methane hydrates represent an enormous carbon and energy source in many low temperature deep marine sediments. However, little information is available concerning the nature of the microbial communities associated with these structures. Here, we describe a phylogenetic analysis based on ribosomal DNA (rDNA) sequences obtained from sediment and fluid samples present in a region of gas hydrate formation in shallow sediments within the Cascadia margin in and around Ocean Drilling Program (ODP) Site 892B. Our studies detected diverse sulfur-utilizing microbes, methanogens, methanotrophs, and non-thermophilic members of the kingdom Crenarchaeota. This is the first culture-independent phylogenetic analysis of a gas hydrate habitat.     

DNA sequence analysis of bacterial toxic heavy metal resistances

Bacterial plasmids have genes that confer highly specific resistances to As, Bi, Cd, Cu, Cr, Hg, Pb, Te, Zn, and other toxic heavy metals. For each toxic cation or anion, generally a different resistance system exists, and these systems may be “linked” together on multiple resistance plasmids. For Cd²⁺, AsO₂ ^?, AsO₄ ^3?, Hg²⁺, and organomercurials, DNA sequence analysis has supplemented direct physiological and biochemical experiments to produce sophisticated understanding. ThecadA ATPase ofS.aureus plasmids is a 727 amino acid membrane ATPase that pumps Cd²⁺ from the cells as rapidly as it is accumulated. This polypeptide is related by sequence to other cation translocating ATPases, including the membrane K⁺ ATPases ofEscherichia coli andStreptococcus faecalis, the H⁺ ATPases of yeast andNeurospora, the Na⁺/K⁺ ATPases of vertebrate animals, and the Ca²⁺ ATPases of rabbit muscle. The conserved residues include the aspartyl residue that is phosphorylated, the lysine involved in ATP binding, and the proline within a membrane translocating region. The arsenate and arsenite translocating ATPase consists of 3 polypeptides (from DNA sequence analysis), including a recognizable ATP binding protein (arsA), an integral membrane protein (arsB gene), and a substrate specificity subunit (arsC gene). Inorganic mercury and organomercurial degradation is carried out by a series of about 6 polypeptides, including 2 soluble intracellular enzymes (organomercurial lyase and mercuric reductase). The latter is related by sequence and function to glutathione reductase and lipoamide dehydrogenase of prokaryotes and eukaryotes. These enzymes are dimeric, FAD-containing, NAD(P)H-dependent oxidoreductases. Other recognizable polypeptides in themer system include a DNA-binding regulatory protein from themerR gene and a Hg²⁺ transport system consisting of a periplasmic Hg²⁺-binding protein (merP gene) and a membrane protein (merT gene) in gram negative systems.