Isolation of the beta-carotene ketolase gene promoter from Haematococcus pluvialis and expression of ble in transgenic Chlamydomonas

摘要

The β-carotene ketolase gene (bkt1) is a key enzyme in the biosynthesis of astaxanthin in the green alga Haematococcus pluvialis. We constructed a genomic library of H. pluvialis from which the upstream sequence of bkt1 was cloned. It was just 27% identical to the β-C-4-oxygenase gene (crto1) promoter. A TATA-box and a number of CAAT-boxes were found in the bkt1 promoter region. Analysis of the sequence revealed the presence of cis-acting elements associated with light and stress-related responses. Seven novel GTAC core sequences involved in copper response were also detected. The bkt1 promoter was transferred into Chlamydomonas reinhardtii CC-849 to drive the expression of ble. The antibiotic resistance and expression of ble in TranBCO transgenic lines confirmed the promoter activity of the cloned bkt1 promoter sequence. The results of this study confirm that the bkt1 promoter owned cis-acting elements involved in light and environmental stresses and the genetic transformation system of C. reinhardtii can be used to study the functions of bkt1promoters from H. pluvialis.