Tobacco plastid transformation using the feedback-insensitive anthranilate synthase [alpha]-subunit of tobacco (ASA2) as a new selectable marker.

Full Abstract

Genetic engineering of chloroplasts normally requires the stable introduction of bacterial derived antibiotic or herbicide-resistance genes as selective markers. Ecological and health concerns have been raised due to the presence of such genes within the environment or the food supply. One way to overcome this issue is the use of plant genes able to confer a metabolic or developmental advantage to the transformed cells manipulating the plant's biosynthetic pathways. We explored the feasibility of using, for plastid transformation, the selection system based on the feedback-insensitive anthranilate synthase (AS) alpha-subunit gene of tobacco (ASA2) as a new selective marker and the indole analogue 4-methylindole (4MI) or the tryptophan analogue 7-methyl-DL-tryptophan (7MT) as the selection agents. An expression cassette containing Prrn-ASA2 was effectively integrated into the region between accD and ycf4 of the tobacco plastome by the biolistic process. Plastid transgenic plants were obtained on medium supplemented with 300 microM 7MT or 4MI. Transplastomic plants showed normal phenotype and fertility and the resistance to the selection agents 7MT and 4MI was transmitted maternally. The plastid transformed lines also exhibited a higher level of AS enzyme activity that was less sensitive to Trp-feedback inhibition and, consequently, increased free Trp levels in leaves about 7-fold.

Author information

Author/s: Barone, Pierluigi (P); Zhang, Xing-Hai (XH); Widholm, Jack M (JM);

Affiliation: University of Illinois, Department of Crop Sciences, Edward R. Madigan Lab, 1201 W Gregory Dr, Urbana, IL 61801, USA. pierluigibarone(-atsign-)gmail.com

Journal and publication information

Publication Type: Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S.

Journal: Journal of experimental botany (J Exp Bot), published in England. (Language: eng)

Reference: 2009-; vol 60 (issue 11) : pp 3195-202

Dates: Created 2009/07/31; Completed 2009/10/19;

PMID: 19553372, status: MEDLINE (last retrieval date: 10/19/2009, IMS Date: )

Sourced from the National Library of Medicine. Abstract text and other information may be subject to copyright.

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