Metabolic engineering of Dunaliella salina for production of ketocarotenoids.
Dunaliella is a commercially important marine alga producing high amount of β-carotene. The use of Dunaliella as a potential transgenic system for the production of recombinant proteins has been recently recognized. The present study reports for the first time the metabolic engineering of carotenoid biosynthesis in Dunaliella salina for ketocarotenoid production. The pathway modification included the introduction of a bkt gene from H. pluvialis encoding β-carotene ketolase (4,4'β-oxygenase) along with chloroplast targeting for the production of ketocarotenoids. The bkt under the control of Dunaliella Rubisco smaller subunit promoter along with its transit peptide sequence was introduced into the alga through standardized Agrobacterium-mediated transformation procedure. The selected transformants were confirmed using GFP and GUS expression, PCR and southern blot analysis. A notable upregulation of the endogenous hydroxylase level of transformants was observed where the BKT expression was higher in nutrient-limiting conditions. Carotenoid analysis of the transformants through HPLC and MS analysis showed the presence of astaxanthin and canthaxanthin with maximum content of 3.5 and 1.9 µg/g DW, respectively. The present study reports the feasibility of using D. salina for the production of ketocarotenoids including astaxanthin.
Department of Botany, St. Xavier's College For Women (Affiliated to Mahatma Gandhi University), Aluva, 683101, India.
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Anila
Plant Cell Biotechnology Department, Central Food Technological Research Institute (A Constituent Laboratory of the Council of Scientific and Industrial Research, CSIR), Mysore, Karnataka, 570 020, India.
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Simon
Bhat Bio-Tech India (P) Ltd., 11-A, 4th Cross Veerasandra Industrial Area Electronics City, Bangalore, 561221, India.
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Chandrashekar
Dr. C. D. Sagar Center for Life Sciences, Dayanada Sagar Institutions, Bangalore, 560008, India.
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Ravishankar
Plant Cell Biotechnology Department, Central Food Technological Research Institute (A Constituent Laboratory of the Council of Scientific and Industrial Research, CSIR), Mysore, Karnataka, 570 020, India. sarada_ravi@yahoo.com.
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Sarada
D015152:Blotting, Northern; D015139:Blotting, Southern; D002338:Carotenoids / Q000096:biosynthesis* / Q000737:chemistry; D000460:Chlorophyta / Q000235:genetics / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D020869:Gene Expression Profiling; D017343:Genes, Plant; D013058:Mass Spectrometry; D060847:Metabolic Engineering / Q000379:methods*; D016133:Polymerase Chain Reaction; D014170:Transformation, Genetic
D002338:Carotenoids
Astaxanthin; Carotenoids; Dunaliella; Metabolic engineering