Polynucleotides encoding thrombin receptor homologs
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[0062] I Isolation of mRNA and Construction of the cDNA Library
[0063] The TRH sequence of this application was first identified in Incyte clone No. 86700 among the sequences comprising the human liver library. The human cells used for this library came from a 49 year old human male (Catalogue # 937224; Stratagene, LaJolla Calif.). cDNA synthesis for the cDNA library was primed with oligo dT, and synthetic adaptor oligonucleotides were ligated onto cDNA ends enabling its insertion into UNIZAP vector system (Stratagene). This allowed high efficiency unidirectional (sense orientation) lambda library construction and the convenience of a plasmid system with blue / white color selection to detect clones with cDNA insertions.
[0064] The cDNA library can be screened with either DNA probes or antibody probes and the PBLUESCRIPT phagemid (Stratagene) can be rapidly excised in vivo. The phagemid allows the use of a plasmid system for easy insert characterization, sequencing, site-directed mutage...
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