An Effective Method for Isolating Genomic DNA from Leaves of Sesame and Perilla

조영수,최용락,정정한,Cho, Young-Su,Choi, Yong-Lark,Chung, Chung-Han 한국응용생명화학회 1997 Applied Biological Chemistry (Appl Biol Chem) Vol.40 No.3

참깨 및 들깨잎으로부터 효율적인 게놈 DNA 분리

정정한,조영수,최용락 ( Chung Han Chung,Young Su Cho,Yong Lark Choi ) 한국응용생명화학회 1997 Applied Biological Chemistry (Appl Biol Chem) Vol.40 No.3

대장균의 저사조절 유전자 nlp의 분자기구 해석

최용락,정수열,정영기,정정한 한국산업미생물학회 1993 한국미생물·생명공학회지 Vol.21 No.3

대장균의 nlp(Ner like protein) 유전자를 클로닝하여 구조해석을 한 결과 전사조절 단백질인 Ner와는 61%의 높은 상동성을 가지고 있음을 이미 보고한 바 있다 . 본 연구는 당 대사 관련 유전자의 발현 조절을 보고자 nlp 유전자를 도입시켜 tac promoter에 의해 대량 발현시킴으로서 lactose 대사 관련 유전자 lacZ와 maltose 대사 관련 유전자인 malQ, malP의 유전자 발현이 2.5배에서 최고 8.3배 정도까지 증가됨을 확인하였다. 이는 nlp 유전자가 cAMP 비존재하에 crp^*1와 상호작용하여 maltose 및 lactose 대사를 촉진시킴을 시사한다. nlp-lacZ 융합 유전자 산물을 Immunoblotting하여 분석한 결과 nlp의 promter가 in vivo 상태에서 발현되어짐을 확인하였다. tac promoter와 IPTG에 의하여 nlp 유전자 산물을 대량 발현시킨 결과 약 10,000 Da의 산물을 SDS-polyacrylamide gel 전기영동으로서 확인하였으며, 부분 정제된 Nlp 단백질을 조절 영역의 DNA 단편에 결합함으로서 전사조절에 관여하는 것으로 사료되어졌다. An nlp (Ner like protein) gene from E. coli was previously cloned and sequenced. Here we show that expression of the sugar metabolism related genes, lacZ, malQ and malP, increased 2.5- to 8.3-fold in the presence of a plasmid containing the nlp gene. This suggested that the nlp gene could induce maltose- and lactose-metabolism coordinately with crp*1 in the absence of cAMP. Using the nlp-lacZ fusion gene, it was possible to show the promoter of nlp was active in vivo. The overexpressed nlp gene product, a polypeptide of 10,000 daltons, was confirmed by SDS-polyacrylamide gel electrophoresis. The band shift assay revealed that the partially purified Nlp protein bound a specific DNA of the regulatory region of the nlp gene.

대장균의 전사조절 유전자 nlp의 분자기구 해석

최용락,정수열,정영기,정정한 東亞大學校附設遺傳工學硏究所 1994 遺傳工學硏究 Vol.- No.1

대장균의 nlp(Ner like protein) 유전자를 클로닝하여 구조해석을 한 결과 전사조절 단백질인 Ner와는 61%의 높은 상동성을 가지고 있음을 이미 보고한 바 있다. 본 연구는 당 대사 관련 유전자의 발현 조절을 보고자 nlp 유전자를 도입시켜 tac promoter에 의해 대량 발현시킴으로서 lactose대사 관련 유전자 lacZ와 maltose 대사 관련 유전자인 malQ, malP의 유전자 발현이 2.5배에서 최고 8.3배 정도까지 증가됨을 확인하였다. 이는 nlp유전자가 cAMP비존재하에 crp*1와 상호작용하여 maltose및 lactose대사를 촉진시킴을 시사한다. nlp-lacZ 융합 유전자 산물을 Immunoblotting하여 분석한 결과 nlp의 promoter가 in vivo 상태에서 발현되어짐을 확인하였다. tac promoter와 IPTG에 의하여 nlp유전자 산물을 대량 발현시킨 결과 약 10,000Da의 산물을 SDS-poly-acrylamide gel 전기영동으로서 확인하였으며, 부분 정제된 Nlp단백질을 조절 영역의 DNA단편에 결합함으로서 전사조절에 관여하는 것으로 사료되어졌다. An nlp (Ner like protein) gene from E. coli was previously cloned and sequenced. Here we show that expression of the sugar metabolism related genes, lacZ, malQ and malP, increased 2.5- to 8.3-fold in the presence of a plasmid containing the nlp gene. This suggested that the nlp gene could induce maltose-and lactose-metabolism coordinately with crp*1 in the absence of CAMP. Using the nlp-lacZ fusion gene, it was possible to show the promoter of nlp was active in vivo. The overexpressed nlp gene product, a polypeptide of 10,000 daltons, was confirmed by SDS-polyacrylamide gel electrophoresis. The band shift assay revealed that the partially purified NIp protein bound a specific DNA of the regulatory region of the nlp gene.

대장균의 전사조절 유전자 nlp의 분자기구 해석

최용락,정수열,정영기,정정한 동의대학교 기초과학연구소 1994 基礎科學硏究論文集 Vol.4 No.1

An nlp (Ner like protein) gene from E. coli was previously cloned and sequenced. Here was show that expression of the sugar metabolism related genes, lacZ, malQ, and malP, increased 2.5- to 8.3-fold in the presence of a plasmid containing the nlp gene. This suggested that the nlp gene could induce maltose- and lactose-metabolism coordinately with crp*1 in the absence of cAMP. Using the nlp-lacZ fusion gene, it was possible to show the promoter of nlp was active in vivo. The overexpressed nlp gene product, a polypeptide of 10,000 daltons, was confirmed by SDS-polyacrylamide gel electrophoresis. The band shift assay revealed that the partially purified Nlp protein bound a specific DNA of the regulatory region of the nlp gene.

An Effective Method for Isolating Genomic DNA from Leaves of Sesame and Perilla

Choi, Yong-Lark,Cho, Young-Su,Chung, Chung-Han 東亞大學校附設遺傳工學硏究所 1998 遺傳工學硏究 Vol.- No.5

Some problems are encountered when isolating genomic DNA from plant materials. These problems are generally derived from co-precipitation of poly saccharides and/or other substances in the genomic DNA preparations Which have great influence on various plant moleculal analyses.^1-4) Until recently, a number of workers have reported a variety of methods for effectively eliminating polysaccharides from plant genomic DNA extractions.^5-9) The general treatments for this involved using high-priced equipment, expensive chemicals or other uncommon materials.^10-13) Some plant materials, for example, yam tissues, grapevine or other wood plant tissues have their own unusual chemical components. Accordingly some appropriate proredures for isolating their genomic DNA were developed, but those procedures were not applicable to the isolation of genomic DNA from leaves of sesame and perilla because mainly of precipitation of unknown substance by applicatien of restrictirin buffers or unacceptable genomic DNA purity. As a result, for isolating functinnal genomic DNA from their leaf tissues, development of a new simple method was needed. Consequently we developed a simple, rapid mothod for isolating genemic DNA from leaf tissues of sesame (Sesamum indicum L.) and perilla (Perilla frutescens) both of whifh could promise as valuable resources for producing useful vegetable oils^17,18) or other commercially important substances.^19,20) In this report, therefore, the procedure that we have developed is described in detail and discussed.

Mutational Analysis of CRP Binding Site in the Regulatory Region of glpD and glpE Genes from E. coli

Choi,Yong-Lark,Chung,Soo-Yeol,Chung,Chung-Han 東亞大學校附設遺傳工學硏究所 1995 遺傳工學硏究 Vol.- No.2

The glpD and glpE genes share a common CRP binding site. Expression of the adjacent but divergently transcribed glpD and glpE genes is positively regulated by the cAMP-CRP complex. In this study, the transcripts of glpD and glpE genes were identified and the positive regulation by cAMP-CRP was also observed at the mRNA level. Several mutations including substitution, deletion, and insertion were introduced into the CRP binding site. We found that several mutants, a 1-bp to 3-bp substitution, an altered distance of the block and have the altered motif, affected the expression of glpD and glpE at different levels. In deletion mutants, which have a single 5´TGTGA3´ motif, the expression of both genes was activated by the cAMP-CRP complex while its effect is greatly reduced. The binding of the cAMP-CRP was reduced by the deletion and substitution mutants.

Molecular Cloning and Sequence Analysis of a New Member of Sialytransferase Gene Family

Lee, Young-Choon,Kim, Hyung-Woon,Chung, Chung-Han,Choi, Yong-Lak,Cho, Young-Su,Kim, Do-Hoon,Kim, Kyung-Sook 東亞大學校附設遺傳工學硏究所 1999 遺傳工學硏究 Vol.- No.6

We have cloned a new member of the sialyltransferase gene family from mouse brain cDNA libraries using sequence information obtained from the conserved amino acid sequence of the previously cloned sialyltransferases. The cDNA sequence included an open reading frame coding for 320 amino acids, and the deduced amino acid sequence showed the highest homology (43% identity) with that of NeuAc α2,3GalB1,3 GalNAcα2,6-sialyltransferase(ST6GalNAcIII) among those of so far cloned mouse sialyltransferases. This open reading frame contained all the characteristic structural features of other sialyltransferases including a type II membrane protein topology and both sialymotifs, one centrally located and the second in the carboxyl-terminal portion of the cDNA. The expression of mRNA was the highest in colon, followed by brain, and low level being found in thymus, lung, heart, spleen, while no signal was detectable in salivery grand, liver and kidney.

Cloning and Characterization of a Seed-Specific ω-3 Fatty Acid Desaturase cDNA from Perilla frutescens

Kim, Joo-Lak,Chung, Chung-Han,Lee, Young-Choon,Choi, Yong-Lark 東亞大學校附設遺傳工學硏究所 1999 遺傳工學硏究 Vol.- No.6

A cDNA encoding ω-3 fatty acid desaturage was isolated from developing perilla seeds and characterized. On the basis of its deduced amino acid sequence comparison, this cDNA was assumed to be a new isoform of microsomal ω-3 fatty acid desaturase gene. Accumulation of the mRNA for this cDNA showed seed-specific expression.

Molecular Cloning and Sequence Analysis of a New Sialyltransferase Gene Family

김경숙,김경운,정정한,최용락,조용수,김도훈,김철호,이영춘 한국생명과학회 1998 한국생명과학회 학술발표회 Vol.20 No.-

We have cloned a new member (STRl) of the sialyltransferase gene family from mouse brain cDNA libraries using sequence information obtained from the conserved amino acid sequence of the previously cloned sialyltransferases. The cDNA sequence included an open reading frame coding for 320 amino acids, and the deduced amino acid sequence showed the highest homology (43% identity) with that of NeuAc α 2,3Galβ1,3GalNAc α2,6-sialyltransferase (ST6GalNAc III) among those of so far cloned mouse sialyltransferases. The primary structure of STRl indicated a type II transmembrane topology, as has been found in all glycosyltransferases cloned to date, consisting of an NH₂-terminal cytoplasmic domain (6 residues), a hydrophobic transmembrane domain (32 residues), and a COOH-terminal active domain (264 residues). The predicted amino acid sequence of STRl contained the highly conserved sialylmotifs L and S, as has been identified in the other cloned sialyltransferases. The expression of mRNA was the highest in colon, followed by brain, and low level being found in thymus, lung, heart, spleen, while signals were not detectable for salivery grand, liver and kidney.