Characterisation and expression of the glutamine synthetase g1n-α gene of french bean

Doctoral thesis English OPEN
Watson, Adam Terence
  • Subject: SB
    mesheuropmc: food and beverages | fungi

Approximately 900bp of the Phaseolus vulgaris cytosolic glutamine synthetase gIn-a gene promoter has been cloned and the DNA sequence determined. The transcriptional start site was mapped using primer extension and RNAse protection techniques. The promoter fragment was fused to the E. coli uidA reporter gene encoding the B-glucuronidase (GUS) enzyme and introduced into transgenic Nicotiana tabacum (tobacco) and Lotus corniculatus. In transgenic tobacco, the gIn-a promoter directed uidA expression in the root-tip, in incipient lateral roots and in emerging lateral roots of all the transgenic plant lines studied. The promoter also directed uidA expression in the root vascular tissue and in the root hairs of a proportion of the lines studied. With the exception of root hairs, the promoter conferred a similar pattern of expression in transgenic L.corniculatus roots and expression of uidA was also observed in senescing mature nodules and possibly in incipient nodules. Expression of gigA was absent from young and mature transgenic nodules. Activity of the gln-a promoter was also associated with several tobacco flower structures including anthers, ovary placenta and vascular tissue, ova, pollen and the stigma and style. Several developing tobacco fruit structures including the placenta, vascular tissue, the developing seed cotyledon and the developing seed coat were also associated with gIn-a. promoter activity. The level of extractable leaf GUS activity was observed to increase between 2- and 5-fold 24 hours after mechanical wounding, relative to non-wounded transgenic leaf tissue. Although twenty four hours after wounding, uidA expression was not associated with those leaf tissues adjacent to the wound site, a higher proportion of wounded leaves showed uidA expression associated with the vascular tissue than in nonwounded leaves. The extractable leaf GUS activity of control cauliflower mosaic virus 358 promoter uidA transgenic plants were seen to decrease up to 50% 24 hours after mechanical wounding relative to non-wounded leaf tissue. Possible physiological functions for gln-a gene expression are discussed.
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