Investigation of function of winged bean promoter in tomato
1994
Napaporn Tantisuwichwong
Medium suitable for the induction of shoot from cotyledon and hypocotyl of 14 - 18 days seedling of tomato was the modified MS medium supplemented with 3 percent sucrose 1 mg/l NAA and 2.5 mg/l BA pH 5.6-5.7. Kanamycin at a concentration of 100 mg/l completely inhibited growth of cotyledon tissue from all three varieties of tomato. Cotyledon tissue was cocultivated for 48 hours with 2*10*[7) - 3*10*[7) cell/ml Agrobacterium tumefaciens LBA 4404 before transferred to the specified medium or modified MS medium supplemented with 3 percent sucrose 1 mg/l zeatin pH 5.0 containing 500 mg/l carbenicillin and 100 mg/l kanamycin. Two sets of constructed genes were used for transferring into tomato. The beta-glucuronidase (GUS) gene was used as a reporter gene. The difference between the two gene contructions was the promoter, one was from a CaMV 35S promoter and the other was from a chymotrypsin inhibitor gene promoter of winged bean. Under the control of CaMV 35S promoter, the expression in transformed tomato was still heterogeneous which supposed to be chimeric plants. Expression of GUS gene under the control of WCI promoter was also heterogeneous but it had a tendency to express in tissue specific manner in immatured seeds of tomatoes as in winged bean. Furthermore the 496 base pairs fragment from GUS gene could be amplified by PCR technique. Therefore the confirmation of transgenic tomato was proved.
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