Molecular cloning and sequence analysis of cDNA encoding human EGF receptor extracellular domain

Aim: To clone the cDNA of human EGFR extracellular domain and to construct the mammalian cell expression vector for soluble EGFR (sEGFR). Methods: The cDNA encoding EGFR extracellular domain was cloned by RT-PCR from human placenta and its sequence was determined. The DNA fragment was inserted into pEGFP-N1 to construct an expression vector for sEGFR. Results: The cDNA encoding the extracellular domain of EGFR was cloned from human placenta villi and the expression vector for sEGFR was constructed by PCR using a primer pair to introduce a Kozak sequence before ATG and to insert a stop codon. The protein encoded by this gene contained domains L1, S1, L2, S2 as well as signal peptide. Sequence analysis revealed that there were 3 nucleotides different from the previously published gene, with G1562→A, G1620→C and T1887→A, respectively. The former two nucleotide substitutions led to amino acid changes at position 497 and 516 from Arg and Lys to Lys and Asn respectively, whereas the latter one was a synonymous mutation (Thr605). Conclusion: The cDNA encoding the extracellular domain of EGFR was successfully cloned and the expression vector for sEGFR was constructed.