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Isolation and characterisation of tilapia β-actin promoter and comparison of its activity with carp β-actin promoter

2003; Elsevier BV; Volume: 1625; Issue: 1 Linguagem: Inglês

10.1016/s0167-4781(02)00534-1

1879-2634

Gyu-Lin Hwang, Md. Azizur Rahman, Shaharudin Abdul Razak, Frédéric Sohm, Hamid Farahmand, Alan M. Smith, C. J. Brooks, Norman Maclean,

RNA Interference and Gene Delivery

The regulatory sequence including proximal promoter, untranslated exon 1 and intron 1 of the β-actin gene from tilapia (Oreochromis niloticus) has been isolated and spliced to a β-galactosidase reporter gene to test its activity. Comparisons of promoter activity have been carried out with three different constructs: (1) 1.6 kb tilapia β-actin regulatory sequence, (2) 1.5 kb carp β-actin regulatory sequence, and (3) 4.7 kb carp β-actin regulatory sequence. Although the 1.6 kb tilapia β-actin regulatory sequence gave slightly different expression patterns in tilapia embryos assayed by in situ X-gal staining, no difference was observed in expression level when the tilapia sequence was compared with the 4.7 kb carp β-actin regulatory sequence by quantitative assay. In comparison with the 1.5 kb carp β-actin regulatory sequence, the 1.6 kb tilapia β-actin regulatory sequence gave higher expression levels in tilapia embryos, while a reverse result was observed in zebrafish embryos. In cell transfection experiments, the 1.6 kb tilapia β-actin regulatory sequence showed three to four times better activity in blue gill cells than either the 4.7 kb carp β-actin or the 1.5 kb carp β-actin regulatory sequences. The 1.6 kb tilapia β-actin regulatory sequence also drove higher reporter gene activity in somatic cells of tilapia than did the 4.7 kb carp β-actin regulatory sequence following direct injection of constructs into muscle. Therefore, taken together, the data demonstrate that the tilapia β-actin promoter can be used as an efficient regulatory sequence to produce autotransgenic tilapia.