New Researches in Biotechnology - Facultatea de Biotehnologii ...

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Proceeding of the 4 rd International Symposium“NEW RESEARCH IN BIOTECHNOLOGY” USAMV Bucharest, Romania, 2011Increase in Bt cotton areaPercent Area90807060504030201009080706050403020100Percent Area-10-102005 2007 2009 2011YearsFigure 1: Evolution of Bt cotton area in Punjab 2006−2010162. MATERIALS AND METHODSNine cultivars of Gossypium Hirsutum L. containing cry1Ac gene (MON531 event) weregrown in RCBD design. The varieties were Ali Akbar 802, IR 3701, Ali Akbar 703,FH113,IR 1524,GH 2035,FH 114,N 121 and MG 6. The plot size was 40 x 20 ft keeping rowto row distance 30 inches and plant to plant distance 12 inches. The sowing date was May15, 2010.Water and fertilizer were applied as per normal recommendations.Three consecutive plants were selected from each genotype in every repeat and tagged after30 days of germination. Selected plants were tested for their Bt studies. Agdiaimmunostrips were used for Bt testing. Cry1Ac protein concentration was quantified incotton leaves by ELISA. After 30 days of sowing, leaf samples were collected andpreserved in liquid nitrogen for ELISA test. Third leaf from the top of each tagged plantwas tested. Tagged plants were quantified five times i.e. 30 days after planting (DAP),70DAP, 100DAP, 130DAP and 160 DAP.Average temperature of days since sowing/previous sampling date was calculated prior toeach sampling. Bt quantification data was analysed by split plot design by using QI Macros2011 software which showed a non normal data nature. At α=0.05, the data rejects nullhypothesis.
Proceeding of the 4 rd International Symposium“NEW RESEARCH IN BIOTECHNOLOGY” USAMV Bucharest, Romania, 20113. RESULTS AND DISCUSSIONThe quantities of Bt toxin determined over the growing season are expressed below (table1).Table 1: Variation in the quantity of Bt toxin over the growing seasonMeanTemperature37 0 C 39 0 C 36 0 C 31 0 C 26 0 CVarieties 30DAP 70 DAP 100 DAP 130 DAP160DAPAA802 0.921 0.755 0.672 0.546 0.541MG6 0.915 0.764 0.722 0.548 0.514IR3701 0.924 0.76 0.681 0.546 0.523AA703 0.92 0.749 0.716 0.548 0.521FH113 0.924 0.765 0.727 0.546 0.527IR1524 0.913 0.756 0.657 0.549 0.506GH2035 0.912 0.765 0.673 0.543 0.504FH114 0.928 0.744 0.636 0.544 0.519N121 0.924 0.743 0.634 0.548 0.516The trend of change in concentration of Bt toxin related with temperature shows the highestproductivity at 37 0 C which declines onwards with an increase in temperature (figure2).The Bt toxin concentration is expressed in µg/g.Bt toxin concentration ranged between0.930—0.510 µg/g. The maximum level of Bt protein was produced at 30DAP whileminimum being at 160 DAP. The concentration of Bt content is tapered off towards the endof the growing period.At 37 0 C, i.e. at 30DAP, the varieties produced the highest quantity of toxin.The juveniletissues produced the maximum effect.However, at this stage the bollworms do not infest thefield as they appear later in the crop season at about 80-100 DAP. At 26 0 C (160DAP), theplants had the favorable temperature for growth and mature plants produced Bt toxinsufficiently. However, the graph clearly indicates that at no growth stage, no plant variety atany temperature range produced Bt toxin high enough to cause 95% mortality of targetpests (high dose strategy).Illegal varieties produced by random crossing of Bollard varietieswith local high yielding cultivars not only cause an economic loss to farmers (as the Btseeds are priced higher than non Bt); but also cause the survival of target pest individuals.The resistance evolution case has not yet been observed in Pakistani cotton fields as noresearch report indicates so. Pakistani cotton experts are of the view that the resistanceevolution in target pests due to less-than-optimal toxin concentration is not feasible(Sharma, 2009). The farmers have small holdings and due to adjacent fields of differentcrop plants, target pests do not survive due to host unavailability in vicinity. However, inmany cotton areas especially nearby cities, there is a trend of vegetable growing nearbycotton fields. The migrating surviving pests may find ochra, brinjal and cauliflower asalternate hosts for development.17
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