Synthesis of the human mature interleukin lα gene by means of the in vitro amplification of the mRNA-cDNA duplex
E. N. Lebedenko, O. V. Plutalov, Yu. A. Berlin
M. M. Shemyakin Institute of Bioorganic Chemistry, Academy of Sciences of the USSR, Moscow
Abstract: Mixture of polyadenylated mRNAs from human monocytes has been subjected to the reverse transcription specifically initiated from the mRNA encoding interleukin la by a synthetic polynucleotide complementary to the mRNA's coding 3'-end to yield the corresponding mRNA-cDNA duplex. Under conditions of the polymerase chain reaction, with the above polynucleotide as a downstream primer and an upstream primer corresponding to the beginning of the mature interleukin 1α (AA 113–271) gene, the mRNA-cDNA duplex yielded the desired gene, whose structure was proved by the restriction and sequence analyses. The gene, containing translation initiation and termination triplets, can be used for producing interleukin lα in various expression systems and as a probe in studies of the lymphokine's biosynthesis. This method of the gene synthesis does not need construction and analysis of cDNA libraries nor synthesis of double-stranded DNA, and can, in principle, make use of the total (non-fractionated) cellular RNA.
Russian Journal of Bioorganic Chemistry 1990, 16 (11):1570-1573