Transformation Of BADH Gene Into Populus×euramericana 'Neva' And Construction Of A Multi-Gene Vector Expressing Enzymes Synthesizing GlyBetaine

Glycine betaine is a very effective osmotic regulator. In higher plant, betaine is synthesized by a two-step reaction: choline→betaine aldehyde→betaine. The first reaction is catalysed by choline monooxygenase (CMO) while the following is catalysed by betaine aldehyde dehydrogenase (BADH). Researches demonstrated that the salt tolerance of BADH gene engineered plants had increased. So far, there is no report concerned populus.In this research Betaine aldehyde dehydrogenase (BADH) gene was transferred into Populus×euramericana 'Neva' by Agrobacterium-mediated leaf disc transformation in order to enhance the salt tolerance. Twenty-five buds survived after antibiotic screening of 650 leaf discs, the following PCR analysis proved that the BADH gene had been integrated into plant genome in 10 of the 25 buds. The salt tolerance test suggested that the transgenic buds grew better in root inducing medium containing 50, 75, 100 mmol/L NaCl in compare with non-transformed buds. In addition, the transgenic plants showed better integrity of cell membrane than wild-type in the same condition of salt stress, which were demonstrated by relative conductivity determination. Chlorophyll determination demonstrated that the transgenic contained more chlorophyll than wild-type.The salt tolerance of plant is attributed to co-operation of several genes, which suggest that a co-expression of betaine synthesizing enzymes in engineered plants would obtain better performance. A multi-gene plant expression vector pYLTAC747H-MAR-BADH-CMO-MAR was obtained by a new method based on Cre/LoxP recombination system and transferred into Agrobacterium Tumefaciens for plant transformation. The choline monooxygenase (CMO) and betaine aldehyde dehydrogenase (BADH) gene derived from Suaeda liaotungensis as well as matrix attachment region (MAR) isolated from tobacco. To simplify the approach, we replaced the electroporation method by heat-shock method for E.coli transformation and eliminated the procedures as dialysis etc.