|
In this work, four F1 hybrids(C1,C2,C8,C10)and one inbred line （2052）were used for anther culture. We related the microspore development stages to the length and shape of buds in tomato. Factors that influence callus induction, such as buds length, disinfection, genotype, medium components and pretreatment, etc., were investigated. In parallel, factors that influence callus propagation such as medium components, concentration of Casein Hydrolysate, Lactoalbumin Hydrolysate, silver nitrate and activated carbon, etc., were studied. At last, factors that influence anther differentiation such as genotype and hormone combination were tested. The aim of all the researches we had done was to optimize a stable and high efficient embryogenic callus induction and plants regeneration system in tomato. Specific results were as follows:1. When the flower bud length was 7.3-10.2 mm, anther length was 4.2-7.1 mm, the corolla and anther were green, yellow-green or yellow, the microspore was in the middle and late uninucleate stage, the induction rate of callus was highest in anther culture. 2. The best disinfection treatment was firstly using 70% alcohol or UV irradiation bud surface 30s, then using 15% NaClO and 2 drops of Tween-20 disinfection l0min.3. The capacity of callus induction was different between different genotypes. Callus induction rate of C10 was highest, which was 34.22%, and was significantly different from the other four genotypes. The best hormone combination to induce callus of anther was 0.5mg.L-1 IAA+1.0mg.L-1 6-BA （MS2） and 0.5mg.L-1 NAA+1.0 mg.L-16-BA （MS5）. Adding 40g.L-1 sucrose can significantly increase callus induction rate. The induction rate of 4℃cold pretreatment for 2 days significantly was 7.78%, which was twice more than the CK. Mean while, the induction rate of 0.6 mol.L-1 mannitol pretreatment for 2 days was 8.14%, which was twice more than the CK.4. The optimum medium for callus propagation was MS+1.00mg·L-1 IAA+1.00mg·L-1 6-BA+30g·L-1 sucrose （M2） and MS+0.50mg·L-1 NAA+ 0.50mg·L-1 KT+30g·L-1 sucrose （M3）, and the highest propagation index was 0.61 in C8. Among all the concentrations, 5.00 g·L-1 CH and 0.50g·L-1 LH were more preferred, and the highest propagation index was 1.00. 10μmol.L-1 silver nitrate could reduce browning of the callus and enhance the vitality of the tissue.5. Medium of MS+2mg.L-1 ZT+0.2mg.L-1 NAA （M7） or 2mg.L-1 ZT （M8） could make callus differentiate into adventitious bud. Genotype also significantly affected the differentiation of callus. The adventitious bud only regenerated from C2 in the test materials, besides, C1 was only got a single piece of leaf, and 2052 was only obtained ten adventitious roots.
|