Cloning of the promoter regions of mouse TGF-beta receptor genes by inverse PCR with highly overlapped primers.
In order to isolate promoters of mouse TGF-beta receptor genes, we used inverse PCR with highly overlapped primers corresponding to the 5' sequence of the receptor cDNAs. Nested primer sets only covered a 30- to 40-base region of the sequences. HinfI-digested and self-ligated mouse genomic DNA was used as a PCR template. Only one band for each receptor was seen after PCR. The amplified DNA fragments could direct luciferase production when the luciferase coding sequence was ligated after the fragments. The sequence of the fragment which correspond to the type II receptor showed partial homology with the promoter region of the human TGF-beta type II receptor. Thus, the inverse PCR with highly overlapped primers could be an easy way to isolate the promoter regions of many genes.
Yoshitomo-Nakagawa
K
K
Department of Virology, University of Tokyo, Japan.
Muramatsu
M
M
Sugano
S
S
D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods; D017931:DNA Primers / Q000235:genetics*; D015252:Deoxyribonucleases, Type II Site-Specific / Q000235:genetics; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D011401:Promoter Regions, Genetic; D018125:Receptors, Transforming Growth Factor beta / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics; D012689:Sequence Homology, Nucleic Acid
D017931:DNA Primers; D018125:Receptors, Transforming Growth Factor beta; D011994:Recombinant Proteins; D015252:Deoxyribonucleases, Type II Site-Specific; C032229:GANTC-specific type II deoxyribonucleases