Sesquiterpene synthases from grand fir abies grandis, and...

C - Chemistry – Metallurgy – 12 – N

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C12N 15/60 (2006.01) A01H 5/00 (2006.01) C07K 14/415 (2006.01) C12N 1/00 (2006.01) C12N 5/04 (2006.01) C12N 9/00 (2006.01) C12N 9/88 (2006.01) C12N 15/29 (2006.01) C12N 15/52 (2006.01) C12N 15/74 (2006.01) C12N 15/79 (2006.01) C12N 15/82 (2006.01) C12N 15/84 (2006.01) C12N 5/06 (2006.01)

Patent

CA 2318531

cDNAs encoding E-.alpha.-bisabolene synthase, .delta.-selinene synthase and .gamma.-humulene synthase from Grand Fir (Abies grandis) have been isolated and sequenced, and the corresponding amino acid sequences have been determined. Accordingly, isolated DNA sequences (SEQ ID No:12; SEQ ID No:19 and SEQ ID No:23) are provided which code for the expression of E-.alpha.- bisabolene synthase (SEQ ID No:13), .delta.-selinene synthase (SEQ ID No:20) and .gamma.-humulene synthase (SEQ ID No:24), respectively, from Grand Fir (Abies grandis). In other aspects, replicable recombinant cloning vehicles are provided which code for E-.alpha.-bisabolene synthase, .delta.-selinene synthase and .gamma.-humulene synthase, or for a base sequence sufficiently complementary to at least a portion of E-.alpha.-bisabolene synthase, .delta.- selinene synthase or .gamma.-humulene synthase DNA or RNA to enable hybridization therewith. In yet other aspects, modified host cells are provided that have been transformed, transfected, infected and/or injected with a recombinant cloning vehicle and/or DNA sequence encoding E-.alpha.- bisabolene synthase, .delta.-selinene synthase or .gamma.-humulene synthase. Thus, systems and methods are provided for the recombinant expression of the aforementioned recombinant sesquiterpene synthases that may be used to facilitate their production, isolation and purification in significant amounts. Recombinant E-.alpha.-bisabolene synthase, .delta.-selinene synthase and .gamma.-humulene synthase may be used to obtain expression or enhanced expression of E-.alpha.-bisabolene synthase, .delta.-selinene synthase and .gamma.-humulene synthase in plants in order to enhance the production of sesquiterpenoids, or may be otherwise employed for the regulation or expression of E-.alpha.-bisabolene synthase, .delta.-selinene synthase and .gamma.-humulene synthase, or the production of their products.

La présente invention concerne des ADN complémentaires codant la E-.alpha.-bisabolène synthase, la .delta.-selinène synthase et la .gamma.-humulène synthase tirées du sapin grandissime (Abies grandis) qui été isolés et séquencés, et les séquences d'acide aminé correspondantes ont été déterminées. De même, cette invention concerne des séquences d'ADN isolées (SEQ ID No:12; SEQ ID No:19 et SEQ ID No:23) codant l'expression de la E-.alpha.-bisabolène synthase (SEQ ID No:13), la .delta.-selinène synthase (SEQ ID No:20) et la .gamma.-humulène synthase (SEQ ID No:24), obtenues à partir du sapin grandissime (Abies grandis). Dans certains aspects de l'invention, des véhicules de clonage recombinés et réplicables codant la E-.alpha.-bisabolène synthase, la .delta.-selinène synthase et la .gamma.-humulène synthase, ou une séquence de base suffisamment complémentaire pour au moins une partie de l'ADN ou ARN de la E-.alpha.-bisabolène synthase, la .delta.-selinène synthase ou la .gamma.-humulène synthase, pour permettre une hybridation. Dans d'autres aspects encore, l'invention concerne des cellules hôtes modifiées ayant été transformées, transfectées, infectées et/ou injectées par le véhicule de clonage recombiné et/ou une séquence d'ADN codant la E-.alpha.-bisabolène synthase, la .delta.-selinène synthase ou la .gamma.-humulène synthase. Par ailleurs, cette invention concerne des systèmes et des procédés d'expression recombinée des sesquiterpène synthases recombinées précitées, pouvant être utilisés pour faciliter leur production, isolement et purification en quantités importantes. Les E-.alpha.-bisabolène synthase, .delta.-selinène synthase et .gamma.-humulène synthase recombinées peuvent être utilisées pour obtenir l'expression ou l'expression accrue de la E-.alpha.-bisabolène synthase, la .delta.-selinène synthase et la .gamma.-humulène synthase dans des plantes, de façon à augmenter la production des sesquiterpénoïdes, ou pour réguler ou exprimer la E-.alpha.-bisabolène synthase, la .delta.-selinène synthase et la .gamma.-humulène synthase, ou encore pour obtenir leurs produits.

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