• BMB Reports
• /
• 제47권2호
• /
• pp.92-97
• /
• 2014

We have examined the effect of bithorax complex genes on the expression of castor gene. During the embryonic stages 12-15, both Ultrabithorax and abdominal-A regulated the castor gene expression negatively, whereas Abdominal-B showed a positive correlation with the castor gene expression according to real-time PCR. To investigate whether ABD-B protein directly interacts with the castor gene, electrophoretic mobility shift assays were performed using the recombinant ABD-B homeodomain and oligonucleotides, which are located within the region 10 kb upstream of the castor gene. The results show that ABD-B protein directly binds to the castor gene specifically. ABD-B binds more strongly to oligonucleotides containing two 5'-TTAT-3' canonical core motifs than the probe containing the 5'-TTAC-3' motif. In addition, the sequences flanking the core motif are also involved in the protein-DNA interaction. The results demonstrate the importance of HD for direct binding to target sequences to regulate the expression level of the target genes.

• Biotechnology and Bioprocess Engineering:BBE
• /
• 제5권5호
• /
• pp.327-331
• /
• 2000

The application of LFH-PCR(long flanking homology region-PCR) for Bacillus subtilis gene disruption is presented. Without plasmid- or phage-vector construction, only by PCR, based on a DNA sequence retrieved from B. subtilis genome data base, kanamycin resistance gene was inserted into two genes of B. subtilis involved in sporulation, spoIIIE and spoIIIG. The effect of gene disruption on subtilisin expression was examined and the sporulation frequency of two mutants was compared to that of the host strain. For this purpose, only 2 or 3 rounds of PCR were required with 4 primers. We first demonstrated the possibility of LFH-PCR for rapid gene disruption to characterize an unknown functional gene of B. subtilis or other prokaryote in the genomic era.

• 한국식물학회:학술대회논문집
• /
• 한국식물학회 1987년도 식물생명공학 심포지움 논문집 Proceedings of Symposia on Plant Biotechnology
• /
• pp.63-79
• /
• 1987

Light regulates a variety of genes in higher plants. The expression of light-induced plant genes is regulated at the level of transcription via red- light photomorphogenic receptor, phytochrome, as well as unknown blue light photoreceptor(s). Ribulose-5-phosphate carboxylase/oxygenase (Rubisco) small subunit (SSB) and light harvesting chlorophyll a/b (Cab) protein are those of the best understood genes regulated by light. 5'-upstream flanking sequence (- -400) of Rubisco SSB and Cab genes sis known as a light responsive, enhance-like element. It responses to red and blue light in transgenic plant system as a tissue specific manner. Phytochrome gene is also regulated by light. In contrast to most of the light regulated plant genes, it is negatively controlled by red light. Search for the cis- and trans-acting factors responsible for the light signal is in progress to understant photomorphogenesis and development in higher plants.

• International Journal of Industrial Entomology and Biomaterials
• /
• 제39권1호
• /
• pp.22-28
• /
• 2019

We previously isolated 9 clones that show stronger signal compared to Bombyx mori cytoplasmic actin gene (BmA3) by using a dot blot hybridization. In this study, we focused on one clone among these clones which has high amino acid similarity with ${\beta}$-tubulin gene of B. mori. This clone was ubiquitously expressed in all tissues and developmental stage of B. mori. As result of promoter assay using dual luciferase assay system, we found the highest transcription activity region (-750/-1) in the 5'-flanking region of ${\beta}$-tubulin gene, which has about 47 fold more intensive promoter activity than BmA3 promoter. Moreover, the ${\beta}$-tubulin promoter was normally regulated in Bm5, Sf9, and S2 cells. Therefore, we suggest that ${\beta}$-tubulin promoter may be used more powerful and effectively for transgene expression in various insects containing B. mori as a universal promoter.

• 한국어병학회지
• /
• 제8권1호
• /
• pp.37-46
• /
• 1995

H. cunea nuclear polyhedrosis virus (HcNPV) 의 polyhedrin 아미노산 및 polyhedrin gene 의 염기서열을 분석하였다. Polyhedrin 은 SDS-PAGE 상에서 3개의 polypeptide band 가 나타났고 주요 polypeptide 는 약 25 Kd 의 분자량을 갖고 있었다. 또한 polyhedrin 은 17 개의 다른 아미노산으로 구성되어 있었다. HcNPV DNA를 EcoRI 효소로 절단하여 $\alpha^{32}P$로 labelling 된 Autographa californica (AcNPV) polyhedrin gene cDNA 의 probe DNA를 이용하여 hybridization 한 결과 polyhedrin gene은 EcoRI 절편들중 H 절편에 양성반응을 나타냈다. 또한 polyhedrin gene 을 포함하고 있는 EcoRI-H 절편을 pUC8 벡터에 cloning한 다음 이를 hPE-H라고 이름하였다. HcNPV genome DNA 의 promoter 부위를 sequence한 결과 TATA box의 염기배열은 polyhedrin gene 전사 개시위치로부터 위쪽으로 -79 bp 의 5' flanking 부위에서 발견되었다. polyhedrin gene 내 CAAT box는 TATA box 측면 염기 배열에서 나타나지 않았고, 4개의 tandem repeat 5'-CTAATAT-3' 와 5'-TAAATAA-3'의 염기는 polyhedrin gene내 전이 개시 위치로 부터 위쪽으로 -141 과 -108 bp 또는 -83 bp 부위에 존재하였으며, 다른 하나는 전이 개시위치로 부터 아래쪽으로 -52 bp 부위에서 발견되었다. 그리고 polyhedrin gene 내 전이 개시위치로 부터 위쪽으로 -141 bp 부위는 다량의 AT (78%) 염기가 존재하였다. 또한 polyhedrin 의 개시 coding region 은 ATG 였고 종결 coding region은 TAA 였다.

• Parasites, Hosts and Diseases
• /
• 제40권4호
• /
• pp.165-172
• /
• 2002

Collaborative studies have identified some genetic factors contributing to the development of severe forms of malaria and schistosomiasis. In Thailand, the $TNF-{\alpha}{\;}5'-flanking$ region shows biallelic polymorphic sites at nucleotides -238, -308, -857, -863, and -1031, and seven alleles have been identified in patients from Myanmar. We found that the TNF promoter (TNFP)-D allele was significantly associated with cerebral malaria in populations from Karen (P < 0.0001. OR ＝ 124.86) and ethnic Burma (P < 0.0001, OR = 34.50) . In China, we have identified two major genes related to the severity of liver fibrosis, one an HLA class II gene, and the other the IL-13 gene. The frequency of the HLA- DRB5*0101 allele and that of the IL-13 promoter A/A (IL- l3P- A/A) genotype were elevated in fibrotic patients, although the two genes are located on different chromosomes, chromosomes 6p and 5q, respectively Subjects with both genotypes had odds ratios (OR = 24.5) much higher than the sum of the ratios for each individual genotype (OR ＝ 5.1,95% Confidence Interval 1.3-24.7 for HLA-DRB5*0101, OR ＝ 3.1 95% CI 1.5 - 6.5 for IL-l3P-A/A). That the effects of the two susceptibility markers are synergistic rather than additive, strongly suggests that the pathogenic Th2 response directly influences the prognosis of post-schistosomal liver fibrosis.

• 생명과학회지
• /
• 제10권4호
• /
• pp.333-339
• /
• 2000

To identify genes involved in the decolorization of crystal violet, we isolated random mutants generated by transponson insertion in crystal violet-declorizing bacterium, Citrobacter sp. The resulting mutant bank yielded mutants with six distinct phenotypes, and Southern hybridization with a Tn5 fragment as a probe showed a single hybridized with six distinct phenotypes, and Southern hybridization with a Tn5 fragment as a probe showed a single hybridized band in the mutants Ctg 2, 5 an 6, whereas two and three bands were detected in Ctg1, 4 and 3, respectively. Tn5-inserted genes were isolated and the DNA sequence flanking Tn5 was determined. From comparison with a sequence database, putative protein product encoded by ctg 5 was identified as E. coli maltose transproter(Mal G) homolog, whereas the deduced amino acid sequence of the other ctg genes did not show any significant similarity with any DNA or protein sequency. Therefore, these results indicate that the other ctg genes except ctg 5 encode new proteins responsible for decolorization of crystal violet.

• Archives of Pharmacal Research
• /
• 제19권6호
• /
• pp.450-455
• /
• 1996

The genomic DNA was prepared from trout liver which was treated with 3-methycholanthrene, and cloned into lambda EMBL3 at BamHl site. The genomic library was constructed via infections of these recombinant phages into E. coli K802, and screened by the most $5^I$-portion of trout CYP1A1 cDNA. After the screening of $10^9$ clones of the amplified library, 12 positive clones were isolated, and subjected to further screenings. The results of southern blot hybridization of genomic DNA prepared from the positive clone showed the presence of a single gene of CYP1A1, and 3.5 Kb PstI fragment that hybridizes with the most $5^I$-region DNA of CYP1A1 cDNA. The restriction map of PstI fragment was determined by the restriction digestion with various enzymes. The nucleotide sequence of the upstream genomic DNA of CYPIAI was determined by DNA sequencing of exonuclease III unidirectionally deleted PstI fragment DNA using $[^{35}/S]$dATP. This paper presented the upstream genomic DNA of CYP1A1 contained a part of coding region which was about 351 base pairs (from ATG to PstI site at 3563).

• Applied Biological Chemistry
• /
• 제39권2호
• /
• pp.104-111
• /
• 1996

감자의 genomic DNA library로 부터 분리한 proteinase inhibitor II (pin2) 유전자들의 제한효소 지도를 작성 하였던바 이미 분리된 상처 유발 (wound-inducible)pin2K 유전자의 것과 상이성이 있는 pin2T를 분리하여 염기서열을 결정하였다. 두 유전자의 염기서열은 전체적으로 약 86%의 동일성을 보였으며 특히 promoter 부위의 염기서열은 pin2K 유전자의 전사개시 부위의 상대적인 위치인 -714까지 네부분의 결손(20 내지 60bp)을 제외하던 약 91%수준의동일성을 보였다. 분리한 유전자들의 promoter 부위를 표지 유전자인 CAT와 GUS 유전자에 연결 시킨후 담배에서의 발현을 추적하였던바, pin2K 유전자의 promoter에 의한 표지유전자의 발현은 상처에 의해 발현 되었으나 pin2T 유전자의 promoter에 의한 표지유전자의 발현은 상처 유무와 관계없이 낮은 수준으로 나타났다. 또한 pin2T 유전자의 Promoter 내의 결손은 핵 단백질의 promoter에의 결합에 영향을 주지 않았으며 상처 유발pin2K 유전자의 promoter 염기서열과 비교하였을때 pin2T 유전자의 promoter 부위내에 5'-AGTAAA-3'라는 특별한 염기부위가 자연적으로 제거된것을 알수 있었다. 또한 5'-AGTAAh-3'의 염기부위가 다른 상처 유발 유전자들에서는 흔히 발견되고, 다른 식물 유전자들의 Promoter에서는 쉽게 발견이 되지 않았다. 따라서 상처 유발 pin2K 유전자의 Promoter내에 상처 유발과 관련있는 특별한 염기부위가 자연적으로 결실되어 pin2T 유전자의 발현이 상처 유발성을 잃은것으로 짐작된다.

• Journal of Microbiology and Biotechnology
• /
• 제1권1호
• /
• pp.45-49
• /
• 1991

The complete nucleotide sequence of alkalophilic Bacillus sp. K-17 $\beta$-xylosidase gene and its flanking regions were established. A 1263-bp of an open reading frame for $\beta$-xylosidase was observed. The molecular weight (50, 521 dalton), deduced from the nucleotide sequence of $\beta$-xylosidase gene, agreed with the result obtained by SDS-polyacrylamide gel electrophoresis of the purified enzyme (51, 000 dalton). The Shine-Dalgarno sequence, 5'-GAGGAGG-3', was found 8 bp upstream of the initiation codon ATG. The -10 sequence (TAAAAT) in the promoter region for $\beta$-xylosidase gene was similar to the consensus sequence for Bacillus subtilis RNA polymerase, whereas the -35 sequence (TCGATCA) different from all the known -35 regions in the promoter for Bacillus subtilis RNA polymerase.