(22)Na influx is significantly lower in salt tolerant groundnut (Arachis hypogaea) varieties.

Distinct varieties differing in salt tolerance were initially identified from two separate green house experiments using two systems; solution as well as soil culture. The first screening involved a diverse group of 27 cultivars. Several physiological traits; Chlorophyll Stability Index (CSI), Salt Tolerance Index (STI) and ion content were determined to screen the cultivars for differences in salt tolerance using solution culture in the first experiment. A set of six varieties (three tolerant and three susceptible) were selected from this experiment and then subjected again to salt stress adopting a natural soil system in the second experiment which involved a screening approach essentially similar to that of the first experiment. In the third experiment using two distinct cultivars differing in salt tolerance selected from experiment II, (22)Na influx rate was determined in the root and shoot at the end of a 24 h salt imposition in Hoagland's nutrient system containing 180 KBq of (22)Na. The results suggested that there were distinct differences in (22)Na influx rate into root and concurrently in the shoot. The salt tolerant Spanish improved and one of the moderately tolerant Trombay variety TAG 24, showed good regulation of (22)Na influx resulting in low (22)Na concentration. The salt susceptible variety JSP39 had nearly 7-8 fold higher root (22)Na content as compared to the tolerant and moderately tolerant cultivars. The results have highlighted the importance of Na exclusion as an important determinant of salt tolerance in groundnut.

Simple yet stringent screening methodologies for evaluation of putative transformants for abiotic stress tolerance: salt and cadmium stress as a paradigm.

Rigorous and stringent screening methodologies to select transformants at both seedling and plant level under cadmium or NaCl stress were developed. At seedling level, two screening strategies were standardized. One involved germination on filter paper/agar in the presence of either CdCl2 (125 µM) or NaCl (350-450 mM) for 9 days and selection of tolerant putative transformants. The other involved germination of the seedlings on soilrite by irrigation of 450 mM NaCl. Further, at plant level, in vitro evaluation for stress tolerance involved a simple leaf senescence bioassay. Combination of the seedling and plant level screening strategies would result in the initial identification of promising transformants for further analysis.

A novel colorimetric assay and activity staining for cytokinin oxidase based on the copper amine oxidase property of the enzyme.

A rapid and sensitive colorimetric assay for cytokinin oxidase (CKO) was developed based on the copper amine oxidase property of the enzyme from cucumber cotyledons. The assay involved oxidation of isopentenyl adenine by CKO in the presence of phenazine methosulphate as an electron acceptor, resulting in reduction of nitroblue tetrazolium to formazon. The formation of formazon was linear with time for at least 20 min and could be followed spectrophotometrically at A490. CKO could also be specifically stained on polyacrylamide gels using the same principle.

Cytokinin oxidase activity and cytokinin content in roots of sunflower under water stress.

Contents of trans-zeatin riboside (ZR), dihydrozeatin riboside (DZR) and N6-(delta2-isopentenyl) adenosine (iPA) was quantified by an indirect ELISA using polyclonal antibodies, in the roots, xylem sap and leaves of pot grown sunflower plants subjected to water stress (RWC of leaves approximately 65 per cent). The delivery rates of all three cytokinins decreased significantly under stress. Cytokinin levels also decreased in roots and in leaves of stressed plants. Three-fold increase in cytokinin oxidase activity was observed in stressed roots after polymin P-ammonium sulphate fractionation. Further purification using Con A agarose resulted in elution of protein with cytokinin oxidase activity and was found to be 30 kDa protein on SDS-PAGE.

Determination of IAA in brassinolide treated coleoptiles of wheat by a modified indirect ELISA with polyclonal antibodies.

An indirect enzyme immunoassay for detection of as little as 10-50 pmole IAA is described for the first time. The assay is based on the development of highly specific polyclonal antibodies against the carboxyl site of IAA. The binding specificity is nearly as high as the radioimmunoassay and the titre of the specific antibody was also remarkably high (1:40,000 of the primary antibody). Such an easy, rapid, specific and highly sensitive assay would be extremely useful in gaining more information on the mode of action of phytohormones, and their effects on physiological processes.

Unusual discrimination against carrier protein antibodies during partial purification of hapten-protein polyclonal antibodies to plant stress hormones.

Polyclonal antibodies (PcAb) were raised against t-zeatin riboside (t-ZR) and abscisic acid (ABA). t-ZR-BSA and ABA-BSA antibodies had high titre and specificity to haptens but also contained BSA specific antibodies as observed in double immuno diffusion and quantitative precipitation tests. Partial purification of antiserum by precipitation, desalting, and ion-exchange chromatography almost completely eliminated interference from BSA specific antibodies. Consequently, very little or no reaction was observed in dot-immunoblot assays even when high concentrations of BSA were probed with partially purified t-ZR-BSA IgG. Further studies with ABA antiserum showed that discrimination against BSA occurred during chromatography and not during salt fractionation. Because antibodies to both hapten and carrier protein were predominantly of IgG class, this unusual discrimination against carrier protein Ab was possibly influenced by two approaches followed for DEAE chromatography, viz. (i) adsorption of IgG at pH 8 followed by elution; or (ii) adsorption of contaminating proteins at neutral pH while specific IgG comes off as unbound fraction.