Identification of promoters of two dehydrogenase genes in Ketogulonicigenium vulgare DSM 4025 and their strength comparison in K. vulgare and Escherichia coli
- 168 Downloads
Promoters of sorbose dehydrogenase gene sdh and sorbosone dehydrogenase gene sndh (Psdh and Psndh) in Ketogulonicigenium vulgare DSM 4025 were identified. The transcription initiation site (TIS) of Psdh was guanine 74 bp upstream of the start codon of sdh and the TIS of Psndh was adenine 113 bp upstream of the first codon of sndh. Comparing Psdh and Psndh, consensus sequences were found, which were TAVCVT (V=A, C or G) and THGAHC (H=A, C or T) for their putative −10 and −35 regions, respectively, and the spans between the 2 regions were 17 bp. Psdh and Psndh promoters may be constitutive in K. vulgare DSM 4025 when cultured in HJ medium. Semiquantitative RT-PCR analysis showed that the Psdh promoter was about 2.5 times stronger than Psndh in strength in K. vulgare DSM 4025. In Escherichia coli, Psdh and Psndh demonstrated strong activity with the former about two times stronger than the latter. DCIP decoloration method and reporter plasmids pSDH or pSNDH may be applied to discover promoters of genes in E. coli and to determine their strength in one step.
KeywordsPromoters Determination Strength comparison Ketogulonicigenium vulgare DSM 4025 E. coli
We are grateful to Professor Susheng Yang for providing vector pBBR1MCS2. This work is supported by the National Natural Science Foundation of China (No. 30470058).
- Sambrook J, Russell DW (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New YorkGoogle Scholar
- Zhang WC, Jiao YH, Yuan HJ, Xie L (2003) A novel l-sorbose dehydrogenase gene and the encoding protein. China Patent, CN03102060.7Google Scholar