Expression of the Volvox gene encoding nitrate reductase: mutation-dependent activation of cryptic splice sites and intron-enhanced gene expression from a cDNA.

摘要: Use of the nitrate reductase encoding gene (nitA) as selection marker has facilitated successful nuclear transformation Volvox carteri. The nitA contains 10 introns. A stable mutation in recipient strain 153–81 resides a G-to-A transition first nucleotide 5′ splice site intron 2. This resulted at least three non-functional variants, namely: (1) 2 was not spliced all; (2) cryptic 60 nt upstream or (3) 16 downstream were activated and used for splicing. When we cDNA (pVcNR13) V. carteri 153–81, low efficiency about 5×10-5 transformants per reproductive cell observed. Re-integration either 1 (pVcNR15) introns 9 (pVcNR16) transforming increased rates to 5×10-4. In parallel, pVcNR15-transformed exhibited growth that 100-fold over pVcNR13-transformed alga. intron-enhancement expression appears be associated with post-transcriptional processing ‘channelling’ message. These data suggest an important role splicing