Cloning and analysis of the full-length Rubisco large subunit (rbcL) cDNA from Ulva linza (Chlorophyceae, Chlorophyta)

摘要

Rubisco (Ribulose-1,5-bisphosphate carboxylase/oxygenase) is the first key enzyme in the photosynthesis pathway. This is the first study to obtain the full-length cDNA encoding Rubisco large subunit (RbsL) from Ulva linza. The partial rbcL cDNA sequence of 1101 bp was first cloned by RTPCR from the total RNA. Both sequences of the 3' and 5' ends were then amplified using 3'-RACE and 5'-RACE technologies, which yielded 605 bp and 371 bp DNA fragments, respectively. A full-length cDNA sequence of 1498 bp was deduced based on the overlapping sequences and was assigned to a new NCBI accession number: DQ813496. It contains a 1425-bp coding region (474 amino acids) for the Rubisco large subunit (RbsL), a leader sequence of 47 bp at 59-UTR, and a polyA site at 3'-UTR. The leader sequence has no Shine-Dalgarno (SD) sequence and was similar to that of Euglena gracilis rbcL with 76.60% A/T content. Bioinformatics analysis and a BLAST search result suggested that the cDNA sequence of U. linza was highly homologous with known full-length cDNAs of the same gene from other green algal species. Three catalytic sites and a CO2 activator region were identified in the Rubisco large subunit of U. linza, and they were completely identical to those of Chlamydomonas reinhardtii and highly conserved with those of spinach and maize. Based on the cDNA sequence, the secondary and tertiary structures of the U. linza Rubisco large subunit were also predicted.