AAB Bioflux, 2014, Volume 6, Issue 2. http://www.aab.bioflux.com.ro 186 AAB BI OFLUX Advances in Agriculture & Botanics- I nternational Journal of the Bioflux Society Effect of proline and salt stress on organogenesis, growth, proline and carbohydrate content of regenerated plantlets in Citrus sinensis (L.) Osbeck cv. Local orange Muayed F. Abbas, Abbas M. Jasim, Huda A. Al-Taha College of Agriculture, Basrah University, Basrah, Iraq. Corresponding author: M. F. Abbas, muayedfadhil@yahoo.co.uk Abstract . The effect of exogenous proline and NaCl stress on organogenesis, growth and certain chemical constituents (soluble carbohydrates and free proline) in Citrus sinensis (L.) Osbeck cv. Local orange in vitro was investigated. Callus cultures of nucleus tissues derived from under developed ovule of immature fruits were used. Primary callus were cultured on different levels of NaCl in the culture media (0, 10, 20, 30, 40 and 50 mM). For interaction experiments, proline was used at 0, 25, 50 and 75 mg L -1 , whereas NaCl concentrations were 0, 40 and 50 mM. Organogenic callus was obtained, when this primary callus was cultured on MS medium supplemented with 5.0 mg L -1 BA, adventitious shoots were obtained when the organogenic callus was incubated on MS medium supplemented with BA at 1.0 mg L -1 and NAA at 0.1 mg L -1 . The addition of NaCl at 10 mM significantly increased the number and length of adventitious shoots, but high concentration of NaCl significantly reduced the number and length, this effect was alleviated by the exogenous application of proline, in particular at 25mg L -1 . Complete plantlets were developed when the adventitious shoots were transferred to half strength MS medium supplemented with 0.1mg L -1 NAA. The addition of NaCl at 10 mM significantly increased plantlet height, leaf number per plantlet, fresh weight of the shoot and root system. However at higher concentration (20 – 50 mM), NaCl significantly decreased all the growth parameters of the plantlets. Proline on its own significantly increased organogenesis, and growth parameters of the plantlets. NaCl significantly decreased soluble carbohydrates of the plantlets and this effect was alleviated by the addition of proline. Proline alone significantly increased carbohydrate levels. NaCl significantly increased free proline of the regenerated plants, and so does exogenous proline treatment. Salt-tolerant plantlets were successfully regenerated, which were transferred to a potting mix containing sand and peat moss (2:1) and grown for 8 months, with a survival rate of 100 %. Key Words: Soluble carbohydrates, callus cultures, plantlets, salt stress, tissue culture. I ntroduction . Salinity is one of the major abiotic stresses, which adversely affect crop productivity and quality, especially in arid and semi arid regions of the world. The problem of soil salinity is further increasing because of the use of poor quality water for irrigation and poor drainage. Adverse effects of salinity on plant growth may be due to ion cytotoxicity, and osmotic stress (Zhu 2003). Most crop plants are susceptible to salinity even when ECe is less then 3.0 ds m -1 . At these salinity levels, the predominant cause of crop susceptibility appears to be ion toxicity rather then osmotic stress (Chinnusamy et al 2005). Citrus is considered as salt sensitive crop, and the critical level of salinity for vegetative growth is 17 mM NaCl, and productivity of the trees decreased by 50 %, when the salinity level is 80 mM NaCl (Storey & Walker 1999). Plant tissue culture techniques have been used to produce salt tolerance cell lines and plants in several species (Munns & Tester 2008). In citrus, callus cultures have been used by some workers to produce salt tolerant plantlets in Citrus sinensis (L.) Osbeck cv. Shamouti (Ben-Hayyim & Kochba 1983). Furthermore, plants subjected to salinity are known to accumulate many organic compounds at high concentration, such as the amino acid proline. There are several report in the literature which showed, that exogenous application proline to cell and callus cultures under salt stress increased salinity tolerance