Metabolic engineering of Dunaliella salina for production of ketocarotenoids

Dunaliella is a commercially important marine
alga producing high amount of b-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 b-carotene ketolase (4,40b-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 lg/g
DW, respectively. The present study reports the feasibility
of using D. salina for the production of ketocarotenoids
including astaxanthin.