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Biomedical subjects

M Naruto

Publications and source records attributed to M Naruto.

23 records · Page 2Linked to original sources

Molecular cloning of complementary DNA for human medullasin: an inflammatory serine protease in bone marrow cells.

Medullasin, an inflammatory serine protease in bone marrow cells, modifies the functions of natural killer cells, monocytes, and granulocytes. We have cloned a medullasin cDNA from a human acute promyelocytic cell (ML3) cDNA library using oligonucleotide probes synthesized from the information of N-terminal amino acid sequence of natural medullasin. The cDNA contained a long open reading frame encoding 237 amino acid residues beginning from the second amino acid of natural meduallasin. The deduced amino acid sequence of medullasin shows a typical serine protease structure, with 41% homology with pig elastase 1.

Amino Acid Sequence↗

Production of recombinant mouse beta-interferon.

A plasmid was constructed to express a mouse beta-interferon (IFN-beta) in Escherichia coli under the control of the modified tryptophan (trp) promoter. E. coli carrying the plasmid were cultivated in a minijar fermentor and synthesized up to 2.7 X 10(6) IU/ml of antiviral activity. At the end of the cultivation the cells became elongated and curved.

Animals↗

Autonomous replicating sequences from intron of human ras gene in a simian virus 40 T antigen dependent system.

Of the several DNA fragments present in the human lung cancer gene, 1.1 and 2.0 kilobase (kb) fragments corresponding to the intron of this gene were hybridized to a half part of the 27 nucleotides perfect palindrome present in the initiation part of replication in simian virus 40 (SV40) DNA. These two fragments cloned in pBR322 had good template activity, and the initiation of DNA replication started from the region of these fragments in an in vitro system, in which the initiation of DNA replication occurs on cloned DNA containing SV40 origin of DNA replication as described previously. Furthermore, these two clones could replicate autonomously in nuclei of SV40 transformed Cos cells, producing SV40 T antigen constitutively when the clones were transfected into Cos cells. These results show that functional SV40 origin-like sequences are present in human genomes, and they can replicate autonomously within the cells which are producing SV40 T antigen.

Antigens, Polyomavirus Transforming↗

Cloned mouse DNA fragments can replicate in a simian virus 40 T antigen-dependent system in vivo and in vitro.

Mouse liver DNA was cut out with BamHI and cloned into YIp5, which contained the URA3 gene of Saccharomyces cerevisiae in pBR322. Of the several plasmids isolated, two plasmids, pMU65 and pMU111, could transform S. cerevisiae from the URA- to the URA+ phenotype and could replicate autonomously within the transformant, indicating that mouse DNA fragments present in pMU65 or pMU111 contain autonomously replicating sequences (ARS) for replication in S. cerevisiae. Furthermore, to determine the correlation between ARS function in yeast cells and that in much higher organisms, we tried to challenge these plasmids with the simian virus 40 (SV40) DNA replication system. Of the two plasmids tested, the EcoRI-BglII region of pMU65 could be hybridized with a chemically synthesized 13-nucleotide fragment corresponding to the origin region of SV40 DNA. Both pMU65 (the EcoRI-BglII region cloned in pBR322) and its subclone pMU65EB could replicate semiconservatively, and initiation of DNA replication started from the EcoRI-BglII region when the replicating activity of these plasmids was tested in the in vitro SV40 DNA replication system we have established before. Furthermore, pMU65 and pMU65EB could replicate autonomously within monkey Cos cells which produce SV40 T antigen constitutively. These results show that a 2.5-kilobase fragment of the EcoRI-BglII region in pMU65 contains the ARS needed for replication in the SV40 DNA replication system.

Animals↗

Synthesis of oligodeoxyribonucleotides by solid phase phosphotriester method on a reduced scale; preparation of oligonucleotides for improved promoter sequence.

A small reaction vessel, composed of a balled sintered glass filter and a test tube with a stopcock, was designed for the solid phase phosphotriester synthesis of oligodeoxyribonucleotide. Operation of small scale synthesis (nucleoside on resin: 3-6 mumol, activated diester of dimer: 20-30 mumol) was performed under the atmosphere of argon. The yield of each coupling reaction was 60-100%. Twelve short oligonucleotides (6-16mer) were obtained in 19-78% overall yield by this method. These oligomers were prepared as a part of the control region of gene to increase the translation efficiency.

Base Sequence↗