Simultaneous accumulation of astaxanthin and β-carotene in Chlamydomonas reinhardtii by the introduction of foreign β-carotene hydroxylase gene in response to high light stress.

Abstract

Carotenoids are important photosynthetic pigments with many physiological functions, nutritional properties and high commercial value. β-carotene hydroxylase is one of the key enzymes in the carotenoid synthesis pathway of Chlamydomonas reinhardtii for the conversion of β-carotene to astaxanthin. The vector p64DZ containing the β-carotene hydroxylase gene crtZ from Haematococcus pluvialis was transformed into C. reinhardtii CC-503. The transformants were selected by alternate culture in solid-liquid medium containing spectinomycin (100µgmL-1). PCR results indicated that the gene crtZ and aadA were integrated into the genome of C. reinhardtii. RT-PCR analysis showed that the gene crtZ was transcribed in Chlamydomonas transformants. HPLC analysis showed that the content of astaxanthin and β-carotene in cells of C. reinhardtii were simultaneously increased. Under medium light intensity cultivation (60µmolm-2s-1), transgenic C. reinhardtii had an 85.8% increase in β-carotene content compared with the wild type. The content of astaxanthin and β-carotene reached 1.97 ± 0.13mgg-1 fresh cell weight (FCW) and 105.94 ± 5.84µgg-1 FCW, which were increased 18% and 42.4% than the wild type after 6h of high light treatment (200µmolm-2s-1), respectively. Our results indicate the regulatory effect on pigments in C. reinhardtii by β-carotene hydroxylase gene of H. pluvialis, and demonstrate the positive effect of high light stress on pigment accumulation in transgenic C. reinhardtii.