Abdul Razzaq

Interaction of callus selection media and stress duration for in vitro selection of drought tolerant callus of wheat

Callus culture is a novel approach addressing cultured cells as selection units independent of whole plant. Natural variations for drought tolerance existing among cell lines can be exploited in vitro in the presence of suitable concentration of osmoticum and stress duration. The study was aimed to standardized callus selection media and culture duration to select drought tolerant cell line (callus) of wheat. Calli were induced from immature embryos on Murashige and Skoog (MS) based medium supplemented with 4 mg/L of 2,4-dichlorophenoxyacetic acid (2,4-D). Proliferated calli were cultured on various callus selection medias (MS based media + 30 g/L sucrose + 6 g/L agar + 4 mg/L 2,4-D + various levels of polyethylene glycol-600 (PEG-6000) induced osmotic stress including -0.3, -0.6, -0.9 or -1.2 MPa along with control, that is, 0.0 MPa) for two, three or four weeks. Callus health, callus growth rate, callus survival (%) and regeneration (%) declined significantly on media supplemented with higher levels of PEG-6000 induced osmotic stress and with increasing stress duration. Not a single callus could survive nor regenerate on selection media comprising PEG-6000 induced osmotic stress of -1.2 MPa imposed for four weeks. The highest callus growth rate (CGR) was recorded on PEG-6000 free media when calli were cultured for four weeks (-7.32%) on the media supplemented with PEG-6000 induced osmotic stress of -1.2 MPa. Callus selection media comprising PEG-6000 which induced osmotic stress of -0.9 MPa for four weeks was found selective and sub-lethal for wheat calli with 8.63% CGR, 26.62% callus survival and 21.50% regeneration; and seemed ideal for screening drought tolerant somaclonal cell lines of wheat.

Proteomic and physiological analyses of wheat seeds exposed to copper and iron nanoparticles

To elucidate the role of Cu and Fe NPs on the yield of wheat varieties, a gel-free proteomic technique was used. NPs were synthesized and characterized through zeta potential, EDX, and SEM. Spike length, number of grains per spike, and 1000 grain weight were increased in wheat varieties treated with 25ppm Cu and Fe NPs. On treatment with 25ppm Cu and Fe NPs, a total of 58, 121, and 25 proteins were changed in abundance in wheat seeds of galaxy-13, Pakistan-13, and NARC-11, respectively. In galaxy-13, exposure to Cu NPs increased proteins involved in starch degradation and glycolysis. Furthermore, the number of proteins related to starch degradation, glycolysis, and tricarboxylic acid cycle was increased in galaxy-13 on Fe NPs exposure. Proteins related to glycolysis and the tricarboxylic acid cycle was increased in Pakistan-13 and NARC-11 by Fe NPs exposure. The sugar content and SOD activity was increased in wheat seeds treated with Cu and Fe NPs. The Cu content was increased at 25ppm Cu NPs exposure in seeds of wheat varieties. These results suggest that Cu NPs improved stress tolerance in wheat varieties by mediating the process of starch degradation, glycolysis, and tricarboxylic acid cycle through NPs uptake.

Gel-free/label-free proteomic analysis of wheat shoot in stress tolerant varieties under iron nanoparticles exposure.

Iron nanoparticles (Fe NPs) have stimulatory effects on the germination ratio and plant growth of wheat. To elucidate the effects of Fe NPs on shoot of drought tolerant Pakistan-13 and salt tolerant NARC-11, a gel-free/label-free proteomic technique was used. The weights/lengths of seedling, shoot, and root of wheat varieties were increased on 5ppm Fe NPs exposure. The number of proteins related to photosynthesis and protein metabolism was decreased and increased in drought tolerant variety and salt tolerant variety, respectively, treated with Fe NPs compared to untreated plants. Differentially changed proteins in drought tolerant variety and salt tolerant variety were mainly related to photosynthesis. Out of photosynthesis related proteins, light reaction was enhanced in salt tolerant variety compared to drought tolerant variety on Fe NPs exposure. The abundance of ribulose bisphosphate carboxylase/oxygenase small chain in drought tolerant variety was higher than that in salt tolerant variety; however, in salt tolerant variety, it was increased 3 fold by Fe NPs exposure compared to untreated plant. These results suggest that Fe NPs improve the growth of wheat seedling, which might be associated with the increase of protein abundance in photosynthesis in salt tolerant variety.

Water stress effects on biochemical traits and antioxidant activities of wheat (Triticum aestivum L.) under In vitro conditions

ABSTRACT Water stress is one of the major environmental stresses that affect agricultural production worldwide, especially in arid and semi-arid regions. This research investigated the effect of water deficit, induced by PEG-6000 on wheat genotypes (GA-2002, Chakwal-97, Uqab-2000, Chakwal-50 and Wafaq-2001) grown in modified MS medium solution. Osmotic stress caused a more pronounced inhibition in leaf relative water content and leaf membrane stability more sensitive (index in Wafaq-2001 and Uqab-2000) genotypes compared with the tolerant (Chakwal-50, GA-2002 and Chakwal-97) genotypes. Upon dehydration, an incline in proline, total soluble sugar, total soluble protein, superoxide dismutase, peroxidase, catalase and malondialdehyde activity content were evident in all genotypes, especially at osmotic stress of −8 bars. The observed data showed that status of biochemical attributes and antioxidant enzymes could provide a meaningful tool for depicting drought tolerance of wheat genotypes. The present study shows that genotypic differences in drought tolerance could be likely attributed to the ability of wheat plants to induce antioxidant defense under drought conditions. In order to develop genotypes with stable, higher yields in dry farming conditions, it is necessary to characterise genetic resources based on drought adaptation, determine suitable genotypes, and then use them in breeding programmes.

Prevalence of internal parasites in sheep/goats and effective economic de-worming plan at upland Balochistan, Pakistan

Experiments were conducted on sheep and goats during 2006 to 2007 to investigate the prevalence, control and efficacy of anthelmintics against internal parasites at two sites (Loralai and Qila Saifullah) in Balochistan, Pakistan. Faecal samples were collected directly from the rectum of 10 to 20% animals from each flock before the administration of anthelmintics. About 87 to 93% sheep/goats were infested with 10 types of internal parasites. These were Dictyocaulus , Moniezia benideni , Moniezia expensa , Fasciola hepatica , Strongyloides , Nematodirus , Trichostrongylus , Cooperia , Trichuris ovis and Bunostomum . The level of parasitic infestation was mostly between 500 to 2000 eggs per gram (EPG) of faeces. Prior to execution of experiment, more than 80% animals showed high parasitic infectivity (>1000 EPG). Drenching of different anthelmintics such as Nilzan, Ivermectin, Zodec, Bendex, Oxadec and Albensil in infested animals with different durations lowered the internal parasitic level (300 EPG) and also eliminated most of the internal parasites except Dictyocaulus . Albensil was found more effective and economical against internal parasites of sheep and goats. It is concluded that 3 to 4 months interval de-worming with suitable anthelmintics enhance the small ruminants’ productivity. Keywords: Small ruminants, internal parasites, anthelmintics and economic parasitic controlA nitrite reductase gene related to somatic embryogenesis was first cloned from Gossypium hirsutum. The cDNA sequence of the gene, named GhNiR, is 2,257 bp in length, with 254 bp of the 5’ untranslated region and 236 bp of the 3’ untranslated region. The open reading frame is 1,767 bp in length, encoding a deduced amino acid sequence of 588 residues with a molecular weight of 65.722 kDa and an isoelectric point of 7.07. Semi-quantitative RT-PCR analysis showed that the expression level of GhNiR was higher in embryogenic calli and somatic embryoids than in nonembryogenic calli among different somatic embryogenesis stages, and that the level of GhNiR mRNA was also higher in the cultivar with higher somatic embryogenesis ability. The catalytic GhNiR was verified by transformation in E. coli BL21 (DE3) strain with the recombinant expression vector pET-28A-GhNiR. NiR activity assay showed that the crude GhNiR protein had obvious activity to NaNO2 substrate.