公开(公告)号：JPS63258597A

公开(公告)日：1988-10-26

申请号：JP9288187

申请日：1987-04-15

Applicant: AGENCY IND SCIENCE TECHN

Inventor： IWAKURA MASAHIRO ,  KOKUBU TOMOKUNI ,  FURUSAWA KIYOTAKA ,  OHASHI SHINICHI ,  TSUDA KEISHIRO

IPC: C12N15/09 ,  C07K1/16 ,  C07K14/00 ,  C07K14/575 ,  C07K14/655 ,  C07K19/00 ,  C12N9/06 ,  C12N13/00 ,  C12P21/02 ,  C12R1/19

Abstract: PURPOSE:To enable purification of the titled protein from a culture liquid, by passing a cell-free extract liquid of cultured microbial cells through an ion exchange column and gel-filtration column. CONSTITUTION:Microbial cells are collected from cultured product of E.coli C600 (FERM P-9301) containing recombinant plasmid pGIF1 and are disintegrated by ultrasonic radiation. The disintegrated product is subjected to centrifugal separation and the obtained supernatant liquid is purified by ion exchange chromatography and subsequent gel-filtration chromatography. The titled protein having amino acid sequence of formula, i.e. fused protein of dihydrofolic acid reductase and somatostatin is produced by the process.

公开(公告)号：JPS62115287A

公开(公告)日：1987-05-26

申请号：JP25566085

申请日：1985-11-14

Applicant: AGENCY IND SCIENCE TECHN

Inventor： IWAKURA MASAHIRO ,  FURUSAWA KIYOTAKA ,  OHASHI SHINICHI ,  TSUDA KEISHIRO

IPC: C12N15/09 ,  C07H21/04 ,  C07K14/60 ,  C12N1/20 ,  C12N1/21 ,  C12P21/02 ,  C12R1/19

Abstract: PURPOSE:A recombinant plasmid containing a gene to code 1st-29th peptide fragments among bovine growth hormone release factor peptide having secretion adjusting activity of growth hormone. CONSTITUTION:Novel recombinant plasmid pGRF29-28 consisting of 5406 pairs of bases having DNA sequence partially shown by formula II, a recombinant plasmid which can provide a host with ampicillin resistance, codes a peptide wherein a peptide sequence consisting of 6 amino acids of methionine-isoleucine- isoleucine-glutamic acid-glycine-alginine is fused to an amino end side of 1st-29th peptide fragment among bovine growth hormone release factor peptide by cleavage with restriction enzyme BclI and can free a DNA sequence consisting of 107 pairs of bases shown by formula I.

公开(公告)号：JPS61271990A

公开(公告)日：1986-12-02

申请号：JP11369285

申请日：1985-05-27

Applicant: AGENCY IND SCIENCE TECHN

Inventor： IWAKURA MASAHIRO ,  FURUSAWA KIYOTAKA ,  TSUDA KEISHIRO

IPC: C12N15/09 ,  C12N1/20 ,  C12N9/06 ,  C12R1/19

Abstract: PURPOSE:A DNA sequence, consisting of a specific base sequence and containing a gene of an enzyme capable of catalyzing a reaction for reducing dihydrofolic acid and producing tetrahydrofolic acid. CONSTITUTION:A DNA sequence expressed by the formula. The DNA sequence consists of a part coding dihydrofolic acid reductase protein of E. coli (sequence A at the 1- - 477-position), part on the upstream side thereof (sequence B at the -56- - -1-position) and part on the downstream side thereof (sequence C consisting of TAA at the 478- - 480-translation stopping number). The sequences (A) and (C) are the same as the sequence derived from a chromosome DNA of E. coli, and the sequence (B) is derived from a chemically synthesized DNA. The sequence (B) consists of a cleavage site of a restriction enzyme, a site of translation stopping group corresponding to the reading frame of three gene translations and ribosome linking sites for E. coli (DNA sequence of AGGA) and for B. subtilis (DNA sequence of AAAGGAGG).

公开(公告)号：JPH0638786A

公开(公告)日：1994-02-15

申请号：JP21813091

申请日：1991-05-21

Applicant: AGENCY IND SCIENCE TECHN

Inventor： IWAKURA MASAHIRO ,  FURUSAWA KIYOTAKA ,  SAKAI TSUKASA ,  TANAKA YOSHIO

IPC: C12N1/21 ,  C12N15/00 ,  C12N15/09 ,  C12N15/53 ,  C12N15/62 ,  C12P21/00 ,  C12P21/02 ,  C12R1/19

Abstract: PURPOSE:To provide a new manifestation vector useful for the production of a protein containing a heterogenic product fused to the carboxy-terminal of DHFR. CONSTITUTION:The manifestation vector pTP104-4 having the DNA sequence of formula. The vector can be prepared by introducing the terminator region of rrnB gene to the downstream of the DHFR gene of pTP70-1.

公开(公告)号：JPH01252290A

公开(公告)日：1989-10-06

申请号：JP7968188

申请日：1988-03-31

Applicant: AGENCY IND SCIENCE TECHN

Inventor： IWAKURA MASAHIRO ,  KOKUBU TOMOKUNI ,  FURUSAWA KIYOTAKA ,  OHASHI SHINICHI ,  SAKAI TSUKASA ,  TANAKA YOSHIO

IPC: C12N15/09 ,  A61K38/22 ,  A61P23/00 ,  A61P25/04 ,  C07K1/14 ,  C07K1/22 ,  C07K14/00 ,  C07K14/575 ,  C07K14/655 ,  C07K14/70 ,  C07K19/00 ,  C12N1/20 ,  C12N1/21 ,  C12N9/06 ,  C12P21/02 ,  C12R1/19

Abstract: PURPOSE:To obtain a large amount of dihydrofolic acid reductaseleucine enkephalin fused protein, by manifesting a fused gene in which a leucine enkephalin gene is included in a plasmid vector pTP 104-4 with a colon bacillus. CONSTITUTION:A newly recombined plasmid pLEK 1, which is replicated in stable with a colon bacillus, imparting resistance to trimethoprim and resistance to ampicillin to a colon bacillus as a host, having a size of 4207 basic pairs and having a bonded structure of chemically synthesized DNA of 34 basic pairs containing conformations coding BamHI of pTP 104-4, 4173 basic pairs DNA fragments of large side obtained by Sa I abscission and a leucine enkephalin, is produced. A colon bacillus introduced of the plasmid pLEK 1 (FERM BP-1818) produces a large amount of dihydrofolic acid reductase-leucine enkephalin fused protein.

公开(公告)号：JPH01230594A

公开(公告)日：1989-09-14

申请号：JP5717288

申请日：1988-03-10

Applicant: AGENCY IND SCIENCE TECHN

Inventor： FURUSAWA KIYOTAKA ,  SAKAI TSUKASA ,  TANAKA YOSHIO

IPC: C07H19/06 ,  C07H1/00 ,  C07H19/16 ,  C07H23/00

Abstract: PURPOSE:To eliminate protecting groups completely under mild conditions in a shortened time, being very beneficial in nucleic acid synthesis, by allowing a trialkylammonium fluoride to act on specific cyclic silicon-protecting derivatives. CONSTITUTION:(A) Trialkylammonium fluoride is allowed to act on (B) cyclic silicon-protecting groups of formula I (R, R' are alkyl, cycloalkyl, aryl, aralkyl, alkaryl; R'' is H, hydroxyl, protected hydroxyl; B is monovalent residue of nucleic acid base or its derivative to effect hydrolysis whereby the compound of formula II is obtained.

公开(公告)号：JPH01144977A

公开(公告)日：1989-06-07

申请号：JP30215487

申请日：1987-11-30

Applicant: AGENCY IND SCIENCE TECHN

Inventor： IWAKURA MASAHIRO ,  KOKUBU TOMOKUNI ,  FURUSAWA KIYOTAKA ,  OHASHI SHINICHI ,  SAKAI TSUKASA ,  TANAKA YOSHIO

IPC: C07K14/575 ,  C07H21/04 ,  C07K14/00 ,  C07K14/195 ,  C07K14/41 ,  C07K14/655 ,  C07K19/00 ,  C12N1/20 ,  C12N1/21 ,  C12N15/00 ,  C12N15/09 ,  C12P21/02 ,  C12R1/19

Abstract: NEW MATERIAL:Recombinant plasmid pTPGIF2 having a size of 4,660 base pairs and a structure obtained by inserting a DNA having a size of 52 base pairs and containing a sequence coding somatostatin into a BamHI incision site of a plasmid vector pTP70-1 capable of fusing a heteropeptide to a carboxy- terminal of a dihydrofolic acid reductase gene of E.coli. USE:Production of fused protein containing somatostatin. PREPARATION:A somatostatin gene is integrated into a plasmid vector pTP70-1. pTPGIF2 is kept in stable state when introduced into E.coli. C600 strain. The E.coli. C600 strain containing pTPGIF2 is deposited in Fermentation Research Institute as FERM BP-1577.

公开(公告)号：JPS63215693A

公开(公告)日：1988-09-08

申请号：JP4964587

申请日：1987-03-04

Applicant: AGENCY IND SCIENCE TECHN

Inventor： FURUSAWA KIYOTAKA ,  SAKAI TSUKASA ,  TSUDA KEISHIRO

IPC: C07H19/073 ,  C07H19/173 ,  C07H23/00

Abstract: PURPOSE:To obtain the titled compound useful as an intermediate in chemical synthesis of nucleic acids in one process, by reacting a nucleoside with a specific silicon halide in the presence of a silver salt of an acid and subsequently hydrolyzing the resultant reaction product. CONSTITUTION:A nucleoside expressed by formula I [B is (protected) nucleic acid base] is reacted with a silicon halide expressed by the formula R2SiX2 or XSiR'2OSiR'X (X is halogen; R and R' are tert-butyl or isopropyl) (preferably di-tert-butyldichlorosilane or tetraisopropyldichlorosilane) in the presence of a silver salt of an inorganic or organic acid (preferably silver nitrate or silver trifluorosulfonate) and the resultant reaction product is subsequently hydrolyzed to afford the aimed compound formula II or III.

公开(公告)号：JPS61260885A

公开(公告)日：1986-11-19

申请号：JP10087885

申请日：1985-05-13

Applicant: AGENCY IND SCIENCE TECHN

Inventor： IWAKURA MASAHIRO ,  FURUSAWA KIYOTAKA ,  KOKUBU TOMOKUNI ,  TSUDA KEISHIRO

IPC: C12N15/09 ,  C07K14/70 ,  C12N1/20 ,  C12P21/02 ,  C12R1/19

Abstract: PURPOSE:To construct a specific recombinant plasmid having a specific DNA sequence, by making it possible to cut out a DNA sequence coding leucine enkephalin consisting of 28 base pairs by cleavage with a restriction enzyme EcoRI. CONSTITUTION:A novel recombinant plasmid pLEG1 obtained by carrying out sorting out of a plasmid vector and chemical synthesis of a DNA coding leucine enkephalin, and passing the synthesized DNA through processes of integration into a plasmid vector, introduction of the resultant recombinant plasmid into Escherichia coli K12C600 strain, separation of microbial cells having the recombinant plasmid and separation of the plasmid. DNAs expressed by formulas I and II synthesized by the phosphoramidite solid phase synthesis method may be used as the DNA coding the leucine enkephalin.