Comparaison de l’expression homologue du gène vip3Aa16 chez Bacillus thuringiensis sous le contrôle de différents promoteurs
Abstract
Abstract: Bacillus thuringiensis produces the vip3 genes during the vegetative growth phase which are active against lepidopteran larvae. During this study, the vip3Aa16 was cloned under the control of different promotors in order to compare its expression and try to improve it. In fact, vip3Aa16 was cloned under the control of 3 promotors in the same time (the spo promotors and the vegetative promotor Prv) (pHT-spo-Prv-vip3Aa16). The vip3Aa16 was also cloned under the control of only the vegetative promotor Prv (pHT-Prv-vip3Aa16).Western blot analyses showed that Vip3Aa16 protein was detected during the vegetative phase but not during the sporulation phase, proving the expression of vip3Aa16 from the Prv and not from the spo promotors. Bioassays against Ephestia kuehniella revealed that the supernatant of BUPM106 (pHT-Prv-vip3Aa16) and BUPM106 (pHT-spo-Prv-vip3Aa16) strains were toxic and caused 30 % of mortality after 5 days.
Résumé: Bacillus thuringiensis synthétise au cours de la phase végétative, les protéines de types Vip3 actifs sur les lépidoptères. Au cours de ce travail, nous avons cloné le gène vip3Aa16 sous le contrôle de différents promoteurs au vue de comparer l’expression et de chercher à l’améliorer. En effet, vip3Aa16 a été cloné sous le contrôle de trois promoteurs à la fois (les deux promoteurs sporulation-dépendants spo et le promoteur végétatif Prv) (pHT-spo-Prv-vip3Aa16). Nous avons également cloné vip3Aa16 uniquement sous le contrôle de son promoteur végétatif Prv (pHT-Prv-vip3Aa16). L’analyse par Western blot a montré la détection de Vip3Aa16 pendant la phase végétative mais non pas durant la phase de sporulation, témoignant l’expression de vip3Aa16 à partir de Prv et la faible ou l’absence d’expression à partir des promoteurs spo. Les tests de toxicité contre les larves d’Ephestia kuehniella a révélé que le surnageant des cultures des souches BUPM106 (pHT-Prv-vip3Aa16) et BUPM106 (pHT-spo-Prv-vip3Aa16) étaient toxiques et ont causé une mortalité de l’ordre de 30 % au bout de 5 jours.
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