Defence-related biochemical changes by elicitor of malformation pathogen,Fusarium mangiferae,in mango

Malformation of mango (Mangifera indica L.) induced by Fusarium moniliforme var. subglutinans is a plant disease of international importance. The paper reports the downstream defence responses at the initial stage in a susceptible host (cultivar Amrapali) after treatment with biotic (isolated from the pathogen cell wall) (BEL) and abiotic (salicylic acid, SA) elicitors, and inoculation of vegetative buds with the pathogen (IVB). The SA was further tested to induce resistance in field trials. The inoculation and application of elicitors increased β-1, 3 glucanase that causes lysis of fungal hyphae by many folds. Hydrogen peroxide (H₂O₂) (active oxygen species) that induces hypersensitive cell death was reduced to the minimum level after treatment with BEL. The reduction of H₂O₂ in the inoculated vegetative buds was also substantial; however, comparatively less with SA treatment. Consequently, there was no hypersensitive cell death in the malformed mango. Salicylic acid that enhances H₂O₂ content by suppressing H₂O₂-degradation by catalase, increased marginally with the SA treatment and in the IVB, but reduced with the BEL. The reduction of SA in BEL-treated buds concomitantly reduced its H₂O₂ content. The activity of catalase, suppressor of resistance mechanism, was reduced in all the treatments, but the reduction was not enough to arrest H₂O₂-degradation. Magiferin (1, 3, 6, 7-tetrahdroxyxanthone C₂-β-D glucoside), a defence metabolite of mango, increased substantially in all the treatments; maximum with the BEL. A pathogenesis-related (PR) protein of 20 KDa that resists symptom development appeared in all the treatments except the control. But light colour of the spots for the PR-protein indicated low protein accumulation. The maximum accumulation was with the IVB followed by SA and BEL treatments. The amount of total protein reduced considerably in all the treatments. The SA treatment on healthy plants failed to induce defence against malformation. Contrarily, the treatment on malformed seedlings restored normal growth within two months. Hence, SA acted better over the infected plants in presence of the pathogen. Thus, a signal transduction system involving SA and H₂O₂ remained nonfunctional and enough defence chemicals could not be synthesised. Defence genes that produce phenolic and β-1, 3 glucanase, however, became activated and saved the plants from death although could not prevent symptom manifestations.     

Development,cross-species/genera transferability of novel EST-SSR markers and their utility in revealing population structure and genetic diversity in sugarcane

Sugarcane (Saccharum spp. hybrid) with complex polyploid genome requires a large number of informative DNA markers for various applications in genetics and breeding. Despite the great advances in genomic technology, it is observed in several crop species, especially in sugarcane, the availability of molecular tools such as microsatellite markers are limited. Now-a-days EST-SSR markers are preferred to genomic SSR (gSSR) as they represent only the functional part of the genome, which can be easily associated with desired trait. The present study was taken up with a new set of 351 EST-SSRs developed from the 4085 non redundant EST sequences of two Indian sugarcane cultivars. Among these EST-SSRs, TNR containing motifs were predominant with a frequency of 51.6%. Thirty percent EST-SSRs showed homology with annotated protein. A high frequency of SSRs was found in the 5′UTR and in the ORF (about 27%) and a low frequency was observed in the 3′UTR (about 8%). Two hundred twenty-seven EST-SSRs were evaluated, in sugarcane, allied genera of sugarcane and cereals, and 134 of these have revealed polymorphism with a range of PIC value 0.12 to 0.99. The cross transferability rate ranged from 87.0% to 93.4% in Saccharum complex, 80.0% to 87.0% in allied genera, and 76.0% to 80.0% in cereals. Cloning and sequencing of EST-SSR size variant amplicons revealed that the variation in the number of repeat-units was the main source of EST-SSR fragment polymorphism. When 124 sugarcane accessions were analyzed for population structure using model-based approach, seven genetically distinct groups or admixtures thereof were observed in sugarcane. Results of principal coordinate analysis or UPGMA to evaluate genetic relationships delineated also the 124 accessions into seven groups. Thus, a high level of polymorphism adequate genetic diversity and population structure assayed with the EST-SSR markers not only suggested their utility in various applications in genetics and genomics in sugarcane but also enriched the microsatellite marker resources in sugarcane.     

Comparative palynological and wood anatomical studies of Indian Taxus wallichiana Zucc., Cephalotaxus mannii Hook. and Cephalotaxus griffithii Hook.

A comparative study of Taxus wallichiana Zucc., Cephalotaxus mannii Hook. and Cephalotaxus griffithii Hook. of two families Taxaceae and Cephalotaxaceae has been carried out in detail to support the taxonomic existence of two families. The comparative studies have been carried out on the basis of wood anatomy and palynology. The anatomical properties of wood including the tracheids, ray parenchyma, axial parenchyma and number of cross-field pits have been described in detail. The palynological studies include the shape, size and ultrastructure of pollen grains. These studies give a taxonomic support for the recognition of two different genera of families Taxaceae and Cephalotaxaceae which are closely related.     

Proteomic identification of camel seminal plasma: Purification of β-nerve growth factor

The camel seminal plasma contains a diverse array of components including lipids, carbohydrates, peptides, ions and proteins. These are essential for maintaining normal physiology of spermatozoa and are secreted mainly from the prostrate, epidydimis and bulbo-urethral glands of reproductive system. The protein profiles of camel seminal plasma were resolved by two-dimensional gel electrophoresis (2D-PAGE). The majority of the protein was found in acidic regions below pI 7.0 and the 19 brightly stained proteins were identified by MALDI-TOF/MS analysis. On the basis of proteomic profiles, β-nerve growth factor (β-NGF) was purified by ion-exchange and gel filtration chromatography and identified by SDS-PAGE and MALDI-TOF/MS analysis. It was further confirmed by western blotting experiments using rabbit anti-β-NGF primary antibody.     

Design and synthesis of 6,7-dimethoxyquinazoline analogs as multi-targeted ligands for α1- and AII-receptors antagonism

Multiple-targeted ligands can have certain advantages for the management of hypertension which has multiple controls. Molecules with dual bioactivities are available in literature for treating metabolic disorders like diabetes, hypertension and hypercholesterolemia. After scrutinizing the SAR of prazosin-type α₁-blockers and AII-antagonists it was planned to develop dual α₁- and AII-antagonists. Five series of quinazoline derivatives were synthesized and evaluated as dual α₁- and AII-antagonists on rat aortic strips for the blockade of known α₁- and AII-agonist mediated contractions. Many compounds showed balanced activity on both the receptors but compound (22) was found to be the most active derivative having higher antagonistic activity on both the receptors. In the in vivo experiments the chosen compound (22) was slightly less active than prazosin but was found to be equipotent to losartan. These findings shed a new light on the structural requirements for both α₁- as well as AII-receptor antagonists.     

Analysis of methionine synthase reductase polymorphism (A66G) in Indian Muslim population

BACKGROUND AND OBJECTIVES:

Methionine synthase reductase (MTRR) is a vital enzyme of homocysteine/methionine metabolic pathway and is required for the conversion of inactive form of methionine synthase (MTR) to its active form. A clinically important allelic variant of MTRR A66G, with less enzymatic activity is reported with worldwide prevalence rate of ~ 30%. The present study was designed to determine the frequency of MTRR A66G polymorphism in rural Sunni Muslim population of Eastern Uttar Pradesh.

MATERIALS AND METHODS:

Total 56 subjects were analyzed for MTRR A66G polymorphism. A66G mutation analysis was carried out according to the polymerase chain reaction-restriction fragment length polymorphism method of Wilson et al. [1] amplification with MTRR specific primers followed by amplicon digestion with NdeI enzyme was used for the identification of different MTRR genotypes in subjects.

RESULTS AND DISCUSSION:

The AA genotype was found in 5 subjects, AG in 23 subjects, and GG genotype in 28 subjects. Genotype frequencies of AA, AG, and GG were 0.089, 0.41, and 0.5 respectively. The allele frequency of A allele was found to be 0.298 and G allele was 0.705.

CONCLUSION:

It is evident from the present study that the percentage of homozygous genotype GG and frequency of G allele is high in the target Muslim population.     

An Alternative Approach in Gateway® Cloning when the Bacterial Antibiotic Selection Cassettes of the Entry Clone and Destination Vector are the Same

The Gateway^® recombination technology has revolutionized the method of gene cloning for functional analyses and high-throughput ORFeome projects. In general, Gateway cloning is highly efficient because after LR recombination and bacterial transformation, only cells containing the recombinant destination clone are selected on an antibiotic selection plate. However, when the antibiotic resistance gene for bacterial selection is the same in the entry and destination vectors, the direct selection of recombinant destination clones on an antibiotic plate is difficult. Here, we demonstrate an efficient and comprehensive approach to obtain positive destination clones directly on an antibiotic selection plate in this situation. The strategy involves polymerase chain reaction (PCR)-mediated amplification of the entry clone using entry vector-specific primers that bind outside the attL sequences and the subsequent use of this purified PCR product for LR recombination with the destination vector. Our results suggest that cloning of linear DNA fragments into circular destination vectors through LR recombination is an efficient method for inserts up to 7 kb in size. Using this approach, the yield of colony PCR positive destination clones was 100 % for genes of various sizes tested in our experiments.     

Dual activity of pyocyanin from Pseudomonas aeruginosa--antibiotic against phytopathogen and signal molecule for biofilm development by rhizobia

The purified pyocyanin from Pseudomonas aeruginosa TO3 was investigated for its antagonistic activity against Macrophomina phaseolina and as a signaling molecule for development of biofilm by rhizobial strain Ca2. The antagonistic activity of purified pyocyanin, as determined by a dry mass method, showed inhibition of M.?phaseolina. Biofilm formation by strain Ca2 was performed by crystal violet assay. There was an increase in biofilm development by Ca2 with an increase in pyocyanin concentration up to 0.12?nmol·L(-1), followed by a reduction. Using a well-diffusion method, we determined the effect of pyocyanin on disease suppression and biofilm formation by strain Ca2 on radicles of groundnut ( Arachis hypogaea L. ) placed in three concentric whorls on water agar plates. Pyocyanin suppressed disease better at high concentration; however, at lower concentrations increased colony-forming units of Ca2 on radicles of seedlings was observed. A field study in soil infested with M. phaseolina showed that a coinoculant of P.?aeruginosa TO3 and rhizobial strain Ca2 enhanced nodule mass and nitrogenase activity by 264.38% and 269.06%, respectively, over that of the control. This study reports that application of pyocyanin-producing pseudomonads together with rhizobia contributes to the enhancement of nodulation ability and better sustains the growth and productivity of groundnut even in the presence of M.?phaseolina.     

VPAC1 (vasoactive intestinal peptide (VIP) receptor type 1) G protein-coupled receptor mediation of VIP enhancement of murine experimental colitis

Distinct roles of the two T cell G protein-coupled receptors for vasoactive intestinal peptide (VIP), termed VPAC1 and VPAC2, in VIP regulation of autoimmune diseases were investigated in the dextran sodium sulfate (DSS)-induced murine acute colitis model for human inflammatory bowel diseases. In mice lacking VPAC2 (VPAC2-KO), DSS-induced colitis appeared more rapidly with greater weight loss and severe histopathology than in wild-type mice. In contrast, DSS-induced colitis in VPAC1-KO mice was milder than in wild-type mice and VPAC2-KO mice. Tissues affected by colitis showed significantly higher levels of myeloperoxidase, IL-6, IL-1β and MMP-9 in VPAC2-KO mice than wild-type mice, but there were no differences for IL-17, IFN-γ, IL-4, or CCR6. Suppression of VPAC1 signals in VPAC2-KO mice by PKA inhibitors reduced the clinical and histological severity of DSS-induced colitis, as well as tissue levels of IL-6, IL-1β and MMP-9. Thus VIP enhancement of the severity of DSS-induced colitis is mediated solely by VPAC1 receptors.