title: Homologous-restraint polymerase chain reaction: an efficient and rapid protocol to clone multiple homologous genes.
abstract: In this article, we present a novel protocol, called homologous-restraint polymerase chain reaction (HRPCR), for cloning multiple homologous genes. One of the homologous genes was cloned by consensus-degenerate hybrid oligonucleotide (CODEHOP) polymerase chain reaction (PCR) and sequenced. Primers of HRPCR were designed with 20 to 30 nt inverted to the known gene before the 5' end of the CODEHOP primers. The amplification of the known gene was restricted owing to the loop of the PCR product or the incorrect binding of the primers and the template. As a result, only unknown genes could be cloned. This protocol proved to be simple, rapid, and efficient. We applied this protocol to clone the multiple homologous genes of beta-1,4-N,6-O-diacetylmuramidase from the genomic DNA of Streptomyces griseus.
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Jinan, China.
authors.forename: Ming
authors.initials: M
authors.lastname: Chen
authors.forename: Honglei
authors.initials: H
authors.lastname: Liu
authors.forename: Yunfeng
authors.initials: Y
authors.lastname: Bai
authors.forename: Zhenghong
authors.initials: Z
authors.lastname: Zhang
authors.forename: Junjun
authors.initials: J
authors.lastname: Liu
authors.forename: Yuzhen
authors.initials: Y
authors.lastname: Zhang
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry / Q000235:genetics; D008969:Molecular Sequence Data; D009113:Muramidase / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D012988:Soil Microbiology; D013305:Streptomyces griseus / Q000201:enzymology* / Q000235:genetics* / Q000302:isolation & purification
chemical_list: D001426:Bacterial Proteins; D017931:DNA Primers; C020026:N,O-diacetylmuramidase; D009113:Muramidase