Genetic Transformation Of Fungi-resistant Genes Into Apple And Tomato

Improving fungal disease resistance is one of the most important breeding target for apples and tomatoes. Transforming foreign antifungal genes into apple and tomato by gene engineering method, once proved conquiring resistance to pathogenic fungi, the transformed plants can be used as the materials of breeding for disease resistance or resistant cultivars. In present research, PGIP gene, chitinase andβ-1,3-glucanase genes were transformed into 'GaLa' apple and 'Zhongshu No.4' tomato mediated by Agrobacterium tumerfaciens. The main results were as follows:Plant expression vector carrying pWR306-PGIP gene was constructed, identificated and then transformed into Agrobacterim tumefaciens. Subsequently PGIP gene was transformed into apple leaves, tomato leaves and shoot fragments mediated by Agrobacterium tumerfaciens. Five transgenic resistant apple buds and eight transgenic resistant tomato plants were proved by PCR preliminarily that PGIP gene had been integrated into apple and tomato genomes.Chitinase andβ-1,3-glucanase genes plant expression vector-pBLGC was transformed into apple leaves, tomato leaves and shoot fragments by Agrobacterium tumerfaciens, obtained 4 apple resistance buds and 9 tomato positive plants by PCR preliminarily.The reaction conditions for detecting activity of chitinase andβ-1,3-glucosanase in tomato were optimized. The results indicated that the optimal temperature and pH enzyme activity of chitinase were 40℃and 5.0, respectively; the optimal reaction time and colouration time were 60 min and 15 min, respectively; the optimal temperature and pH for enzyme activity ofβ-1,3-glucosanase were 50℃and 4.5 or 8.5, respectively; the optimal reaction time and colouration time were 30 min and 20 min, respectively. Enzyme avtivity assay indicated that enzyme activities in transgenic plants improved a lot than control plants, the highest transgenic chitinase andβ-1,3-glucosanase activities reaches 7.27%~127.27% and 37.63%~279.64%, respectively.The fungi resistance of positive transgenic tomato with chitinase andβ-1,3-glucanase were tested. Atifungal activity of crue chitinase enzyme extraction from the leaves of the transgenic plants to Apple jonathan spot and Grape anthracnose improved 50% and 49.8% than control plants, respectively; antifungal activity of theβ-1,3-glucosanase enzyme crue extraction from the leaves of the transgenic plants to Apple jonathan spot and Grape anthracnose improved 50% and 33.3% than control plants, respectively; antifungal activity of the combinationβ-1,3 -glucosanase and chitinase enzyme extraction from the leaves of the transgenic plants to Apple jonathan spot and Grape anthracnose all improved 66.6% than control plants.To detect the pathogenic bacteria PG resistance of positive transgenic plant, antifungal activity of the crue PGIP enzyme extraction from leaves of the transgenic plants to Aspergillus japonicus PG improved 45% than control plants.