Polypeptide-human SEH protein 29 and polynucleotide encoding this polypeptide
A polynucleotide and protein technology, applied in the field of polynucleotide and polypeptide preparation, can solve the problem of low cyclooxygenase activity
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Embodiment 1
[0086] Example 1: Cloning of human sEH protein 29
[0087] Total RNA was extracted from human fetal brain by one-step method of guanidine isothiocyanate / phenol / chloroform. Poly(A) mRNA was isolated from total RNA using Quik mRNA I isolation Kit (product of Qiegene). 2ug poly(A) mRNA was reverse transcribed to form cDNA. Smart cDNA cloning kit (purchased from Clontech) was used to insert the cDNA fragment into the multiple cloning site of the pBSK(+) vector (product of Clontech Company), transform DH5α, and the bacteria formed a cDNA library. The sequences of the 5' and 3' ends of all clones were determined using Dyeterminate cycle reaction sequencing kit (product of Perkin-Elmer) and ABI 377 automatic sequencer (Perkin-Elmer). Comparing the determined cDNA sequence with the existing public DNA sequence database (Genebank), it was found that the cDNA sequence of one of the clones, 0871b06, was a new DNA. The insert cDNA fragment contained in this clone was determined bidirec...
Embodiment 2
[0088] Example 2: Homology retrieval of cDNA clones
[0089] The sequence of the human sEH protein 29 of the present invention and the protein sequence encoded by it were published in Genbank, Databases such as Swissport conduct homologous searches. The gene with the highest homology to human sEH protein 29 of the present invention is a known human sEH protein, and the accession number of the encoded protein in Genbank is L05779. The results of protein homology are shown in Figure 1. The two are highly homologous, with an identity of 40%. Example 3: Cloning of the gene encoding human sEH protein 29 by RT-PCR
Embodiment 3
[0089] The sequence of the human sEH protein 29 of the present invention and the protein sequence encoded by it were published in Genbank, Databases such as Swissport conduct homologous searches. The gene with the highest homology to human sEH protein 29 of the present invention is a known human sEH protein, and the accession number of the encoded protein in Genbank is L05779. The results of protein homology are shown in Figure 1. The two are highly homologous, with an identity of 40%. Example 3: Cloning of the gene encoding human sEH protein 29 by RT-PCR
[0090] The total RNA of fetal brain cells was used as a template, and oligo-dT was used as a primer to carry out reverse transcription reaction to synthesize cDNA. After purification with a Qiagene kit, PCR amplification was performed with the following primers:
[0091] Primer1: 5'-GGCTCTTCTGGTCCCTGGTCTACTG-3'(3)
[0092] Primer2: 5'-TAATATAACTTTTTAATGGTTAAAA-3'(4)
[0093] Primer1 is the forward sequence starting from...
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