EFFECT OF SELECTED INSECTICIDE ON WHITEFLY (Bemisia tabaci) INFESTING BRINJAL PLANTS 506 In-Vitro Conservation of Sugarcane Germplasm Rabia Ikram, Shazia Erum, Hina Javed, Shah Zaman, Shahid Khan, Aish Muhammad and Saima Kanwal Plant Genetic Resource Institute, NARC, Islamabad Abstract Lack of in vitro multiplication procedure has long been a serious problem in sugarcane breeding programs. This problem has been solved to maximum extent by In Vitro propagation. In the present study, 6 genotypes of sugarcane (SC 06, SC 04, SC 25, SC 11, SC 13 and SC 30) was acquired & excised for apical and sub apical parts (eyes/ lateral buds). The explants were surface sterilized with Clorox (70% +100%, 20 & 10 min). Results showed that highest survival % was achieved with apical buds (66%) as compared to lateral buds (33%). In addition, effect of different concentrations of rooting media (NAA, 5% Sucrose and 6% Sucrose) and shooting media (GA3, 0.1mg/, Kinetin, 0.1mg/L and BAP 0.1mg/L, Kinetin 0.1mg/L,GA3 0.1mg/L, NAA 0.1mg/L) was studied. Synergistic response was observed on the growth of plant with hormone consortia. Significant differences were observed in No. of roots, shoot length @ No. of shoots at all tested treatments. Highest No. of roots recorded for SC 30 at with 6% Sucrose. Highest No. of shoots was recorded for SC 04 with GA3 @ 0.1mg/L and highest shoot length for SC 06 at 0.1mg/ L of Kinetin. Keywords: Growth regulators, in vitro preservation, saccharum officinarum, sugarcane Introduction 1 Conservation of plant genetic resources via tissue culture received immense attention from researchers in the last three decades. Storage of shoot tips or meristem derived explants under slow rate of growth has a significant use in the international germplasm resources units. This procedure makes germplasm available at any times for international distribution. Several types of plant materials have been used for in vitro preservation of clonally propagated crops. Meristem derived explants such as shoot tips and buds were mostly suggested for their genetic and generative stability (Baksha et al., 2002). Sexually propagated plants may be stored as seeds and are not in danger if compared with vegetatively propagated plants. Clonally Corresponding author’s Name: Saima Kanwal Email address: saima.kanwal80@gmail.com propagated plants are usually maintained in botanical gardens. Some of those plants are usually preserved by continuous multiplication of tubers, roots, cuttings or bulbs. Such a procedure is laborious and exposes plants to pests and environmental stresses. Tissue culture has proved to be a useful tool for storage of vegetatively propagated commercial crops like potato, palm, forest trees, fruit trees and other species. Sugarcane is a member of the genus Saccharum from family Gramineae. It is a high valued cash crop and exclusive source of 75% world sugar production (Lakshmanan et al., 2006). This crop provides many by- products for bio-factory such as alcohol, butanol, acetic acid, animal feed and paper besides, sugar and energy (Garcia et al., 2007). Varieties of sugarcane are highly heterogeneous and generally multiplied vegetatively by stem cutting. Lack of suitable multiplication procedure and contamination by systemic diseases are the Asian Journal of Agriculture and Rural Development journal homepage: http://aessweb.com/journal-detail.php?id=5005 mailto:saima.kanwal80@gmail.com Asian Journal of Agriculture and Rural Development, 4(11)2014: 506-509 507 serious problem to multiply an elite genotype of sugarcane in the open field (Lal and Singh, 1994). Tissue culture of sugarcane has received considerable research attention because of its economic importance as a cash crop. Plant regeneration through tissue culture technique would be a viable alternative for improving the quality and production of sugarcane. Material and methods Sugarcane varieties namely SC 06, SC 04, SC 25, SC 11, SC 13 and SC 30 were acquired. Sugarcane in Vitro cultures were established using standard procedure (Uzma et al., 2012). Surface sterilization was done using 70% and 100% Clorox (Commercial bleach comprising 5.25% v/v sodium hypochlorite; an active agent) or 20 and 10 min of time duration under aseptic conditions. After culture establishment, the effect of different concentrations of rooting media (NAA, 5% Sucrose and 6% Sucrose) and shooting media (GA3, 0.1mg/L, Kinetin, 0.1mg/L and BAP 0.1mg/L, Kinetin 0.1mg/L,GA3 0.1mg/L, NAA 0.1mg/L) on rooting and shooting respectively, was studied. The cultures were incubated at 25˚C temperature and 16hr light period. Data for number of leaves, number of roots was recorded after 1 month of culturing. Results and discussion Results showed that highest survival percentage was achieved with apical buds (66%) as compared to lateral buds (33%). %age survival of explants is generally low in case of treatment with 70% Clorox for 10 min. However, when the concentration of Clorox is increased to100%, the survival percentage of explants is elevated 2 to 4 times. In addition, effect of different concentrations of rooting media (NAA, 5% Sucrose and 6% Sucrose) and shooting media (GA3, 0.1mg/, Kinetin, 0.1mg/L and BAP 0.1mg/L, Kinetin 0.1mg/L,GA3 0.1mg/L, NAA 0.1mg/L) was studied. Significant differences were observed in number of roots, shoot length and number of shoots at all tested treatments. (Table 1). The genotype SC 30 exhibited highest number of roots on Treatment 6% Sucrose. Whereas, highest number of shoots (8) were recorded for SC 04 and maximum shoot height (9.667cm) was given by SC 30 at GA3+0.1mg/L. These results indicate that SC 30 was the best performer among all genotypes. These genotypes are now available in disease-free state at the In Vitro Conservation Lab, Plant Genetic Resource Institute (PGRI), NARC which can be used any time for further multiplication, characterization and research. Table 1: Analysis of variance table for rooting Source of variation DF Number of Roots Number of shoots Shoot length Rep 2 6.222 8.722 0.728 Var 5 336.500** 12.388** 41.713** Treat 2 149.733** 0.722 18.623** Var X Treat 10 67.367** 7.677** 9.697** Error 34 3.065 0.741 0.102 Total 53 CV 27.64 29.25 5.27 Table 2: Effect of different rooting media on number of roots Variety X Treatment Number of Roots SC 06 XNAA 4.333 EFGH SC 06 X 5% Sucrose 1.000 H SC 06 X 6% Sucrose 0.333 H SC 04 X NAA 2.333 GH SC 04 X 5% Sucrose 1.000 H SC 04 X 6% Sucrose 18.333 AB SC 25 X NAA 2.000 H SC 25 X 5% Sucrose 1.333 H Asian Journal of Agriculture and Rural Development, 4(11)2014: 506-509 508 SC 25 X 6% Sucrose 8.333 CDEF SC 11 X NAA 2.333 GH SC 11 X 5% Sucrose 10.000 CD SC 11 X 6% Sucrose 7.667 CDEFG SC 13 X NAA 0.667 H SC 13 X 5% Sucrose 3.667 FGH SC 13 X 6% Sucrose 9.667 CDE SC 30 X NAA 5.333 DEFGH SC 30 X 5% Sucrose 13.000 BC SC 30 X 6% Sucrose 22.667 A Table 3: Effect of different shooting media on number of shoots and shoots length Variety X Treatment Number of Shoots Shoot Length SC 06 X GA3, 0.1mg/L 1.3333 D 5.1333 CDE SC 06 X Kinetin, 0.1mg/L 1.6667 CD 8.1667 B SC 06 X BAP 0.1mg/L, Kinetin 0.1mg/L,GA3 0.1mg/L, NAA 0.1mg/L 3.6667 BCD 7.9667 B SC 04 X GA3, 0.1mg/L 8.0000 A 3.8667 FG SC 04 X Kinetin, 0.1mg/L 4.0000 BC 2.5333 H SC 04 X BAP 0.1mg/L, Kinetin 0.1mg/L,GA3 0.1mg/L, NAA 0.1mg/L 2.3333 CD 9.9000 A SC 25 X GA3, 0.1mg/L 1.6667 CD 4.7333 DEFG SC 25 X Kinetin, 0.1mg/L 1.6667 CD 6.0000 C SC 25 X BAP 0.1mg/L, Kinetin 0.1mg/L,GA3 0.1mg/L, NAA 0.1mg/L 2.3333 CD 4.8000 DEF SC 11 X GA3, 0.1mg/L 2.6667 CD 3.7667 G SC 11 X Kinetin, 0.1mg/L 5.6667 AB 4.0000 FG SC 11 X BAP 0.1mg/L, Kinetin 0.1mg/L,GA3 0.1mg/L, NAA 0.1mg/L 3.6667 BCD 5.4667 CD SC 13 X GA3, 0.1mg/L 3.3333 BCD 2.6667 H SC 13 X Kinetin, 0.1mg/L 2.0000 CD 5.9667 C SC 13 X BAP 0.1mg/L, Kinetin 0.1mg/L,GA3 0.1mg/L, NAA 0.1mg/L 2.6667 CD 4.2333 EFG SC 30 X GA3, 0.1mg/L 2.0000 CD 9.9667 A SC 30 X Kinetin, 0.1mg/L 2.3333 CD 9.9000 A SC 30 X BAP 0.1mg/L, Kinetin 0.1mg/L,GA3 0.1mg/L, NAA 0.1mg/L 2.0000 CD 9.9667 A Asian Journal of Agriculture and Rural Development, 4(11)2014: 506-509 509 Figure 1: Performance of genotypes on treatments Conclusions It is concluded from this study that apical buds give better result as compared to lateral buds. Results indicate that SC 30 performed best among all genotypes. These genotypes available at the In Vitro Conservation Lab, Plant Genetic Resource Institute (PGRI), NARC which can be used any time for further multiplication, characterization and research. References Baksha, R., Alam, R., Karim, M. Z., Paul, S. K., Hossain, M. A., Miah, M. A. S & Rahman, A. B. M. M. (2002). In vitro shoot tip culture of sugarcane (Saccharum officinarum) variety LSD 28. Biotechnology, 1(2-4), 67- 72. Garcia, R., Cidade, D., Castellar, A., Lips, A., Magioli, C., Callado, C., & Mansur, E. (2007). In vitro morphogenesis patterns from shoot apices of sugarcane are determined by light and type of growth regulator. Plant Cell Tiss. Org. Cult, 90, 181–190. Lakshmanan, P., Geijskes, R. J., Wang, L. F., Elliott, A., Grof, C. P. L., Berding, N., & Smith, G. R. (2006). Developmental and hormonal regulation of direct shoot organogenesis and somatic embryogenesis in sugarcane (Saccharum spp. interspecific hybrids) leaf culture. Plant Cell Rep, 2, 1007–1015. Lal, N., & Singh, H. N. (1994). Rapid clonal multiplication of sugarcane through tissue culture. Plant Tissue Cult, 4, 1-7. Uzma, M., Khan, R., Muhammad, A., Hussain, I., Shah, S. H., Kumar, T., Inam, S., Zubair, M., Rehman, H. U., Sher, A., Rehman, N., Ahmed, S., & Ali, G. M. (2012). Rapid in vitro multiplication of sugarcane elite genotypes and detection of sugarcane mosaic virus through two steps RT-PCR. Int. J. Agric. Biol, 14, 870‒878. 0 2 4 6 8 10 12 14 16 SHOOT LENGTH (cm) NO. OF SHOOT NO. OF ROOTS SHOOT LENGTH (cm) NO. OF SHOOT NO. OF ROOTS SHOOT LENGTH (cm) NO. OF SHOOT NO. OF ROOTS GA3 0.1 mg/L kinetin 0.1 mgL kinetin 0.1 mgL+GA3 0.1 mgL+ BAP 0.1 mg/L SC-06 SC-04 SC-25 SC-11 SC-13 SC-30 Linear (SC-06)