ISSN:
1573-5028
Keywords:
cryptic splice sites
;
intron-enhancement
;
gene expression
;
nitA cDNA
;
Volvox
;
transformation
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
Notes:
Abstract Use of the nitrate reductase encoding gene (nitA) as selection marker has facilitated the successful nuclear transformation of Volvox carteri. The Volvox nitA gene contains 10 introns. A stable nitA mutation in the Volvox recipient strain 153–81 resides in a G-to-A transition of the first nucleotide in the 5′ splice site of nitA intron 2. This mutation resulted in at least three non-functional splice variants, namely: (1) intron 2 was not spliced at all; (2) a cryptic 5′ splice site 60 nt upstream or (3) a cryptic 5′ splice site 16 nt downstream of the mutation were activated and used for splicing. When we used nitA cDNA (pVcNR13) for transformation of V. carteri 153–81, a low efficiency of about 5×10-5 transformants per reproductive cell was observed. Re-integration of either intron 1 (pVcNR15) or introns 9 and 10 (pVcNR16) in the transforming cDNA increased transformation rates to 5×10-4. In parallel, pVcNR15-transformed Volvox exhibited growth rates that were 100-fold increased over the pVcNR13-transformed alga. This intron-enhancement of nitA gene expression appears to be associated with post-transcriptional processing and ‘channelling’ of the message. These data suggest an important role of splicing for gene expression in V. carteri.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00020601
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