Cloning, Sequencing and
Preliminary Expression of Human RP2 Gene
LIU Xiao-Jun, CHAI Jian-Hua
( Human Genome Lab, Institute of Genetics, Fudan University, Shanghai 200433,
China )
Abstract RP2
is an X-linked retinitis pigmentosa gene, which was newly discovered by
positional cloning. A polymerase chain reaction (PCR) was conducted to screen a
full-length cDNA fragment, defined as hRP2a, which included the coding
region of hRP2, in a human retina cDNA library. hRP2a gene
was cloned into the pJLA503 vector and hRP2 gene was subcloned into
the expression vector pPROEX HTa. Polymorphism was demonstrated at
two sites through DNA sequencing. The recombinant pPRORP2
was transformed into Escherichia coli strain DH5α and the expression
of a 6×His tagged hRP2 fusion protein was induced by IPTG. Band density
scanning of stained gel was performed to estimate the percentage of the
recombinant protein in the total bacterial protein. The ratio was 7% when the
expression was induced at 30 ℃ and was 5.6 % at 37 ℃. The cloning and
expression of hRP2 gene in E.coli established a basis for the
further purification and studies of RP2 for its physiochemical identity,
immunohistochemistry and structure-function relationship.
Key words hRP2； gene cloning； DNA sequencing； gene expression
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