• Fisheries and Aquatic Sciences
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• v.6 no.3
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• pp.116-124
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• 2003

Isolation and cloning of seven-band grouper (Epinephelus septemfasciatus) growth hormone cDNA from pituitary gland revealed an open reading frame of 612 bp coding for a pre-growth hormone of 204 amino acids with a 17 amino acid putative signal peptide. Deduced amino acid sequence showed that there was one possible N-glycosylation site at $Asn^{l84}$ and four cysteine residues $(Cys^{52},\;Cys^{160},\;Cys^{177},\;Cys^{185})$ on t e same positions as in some other species where they were involved in the stabilization of the tertiary structure. The seven-band grouper growth hormone (sbgGH) presented a $99.5\%$ amino acid sequence identity with the growth hormone of Epinephelus coioides and contained the conserved hormone domain region. Comparison of growth hormone sequences from evolutionarily diverse species revealed 25 amino acid residues conserved in jawless fishes to modern mammals. It also revealed an evolutionary trend to retain the same polypeptide sequence even in the distantly related animals while allowing alterations to occur in polypeptides of the closely related species. In order to create a recombinant system to produce high levels of the growth hormone, it was expressed in Escherichia coli (BL21) cells. The gel analysis revealed theoretically expected molecular weights for both mature and pre-sbgGHs.

• Microbiology and Biotechnology Letters
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• v.16 no.4
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• pp.316-319
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• 1988

Pectate Lyase (PL) was cloned from alkali-tolerant Bacillus sp. YA-14 into Escherichia coli MB1000 by inserting HindIII-generated DNA fragment into the HindIII site of pBR322 and then screening recombinant transformant for the ability to hydrolyze sodium polypectate on agar plate, The recombinant plasmid, called pYPC29, was isolated, and the size of the cloned HindIII fragment was found to be 1.6 kb. The PL gene was stablely maintained and expressed efficiently in Escherichia coli. The Pt accumulated largely in the periplasmic space of Escherichia coli clones.

• Journal of Microbiology and Biotechnology
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• v.28 no.7
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• pp.1185-1198
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• 2018

The nucleosomal organization of chromatin using histone proteins is a fundamental and ubiquitous feature of eukaryotic nuclei, with the major exception of dinoflagellates. Although a number of recent genomic and transcriptomic analyses have detected numerous histone genes in dinoflagellates, little is known about their expression. Here in, we aimed to investigate the expression pattern of histone genes under nutritional stress, and an attempt was made to detect histone expression at the protein level in Alexandrium pacificum. The presence of histones at the mRNA level was confirmed in this study by the amplification, cloning, and sequencing of 10 different genes. Relative expression profiling of these genes under different growth conditions was determined with real-time PCR and revealed considerable levels of histone transcription in nutritionally stressed cells. We were unable to detect the expression of histones at the protein level even after immunodetection and analysis using mass spectrometry, although a histone-like protein was detected as a major nuclear component. A. pacificum expresses multiple variants of histone, and protein sequences revealed both conservation and divergence with respect to other eukaryotes. We concluded that A. pacificum maintained an active transcription of histone genes within the cell, and enhanced expression of histone genes in nutritional stress strongly suggest that histones have functional significance in dinoflagellates, although expression at the protein level was below our current detection limits, which suggests a limited role of histones in DNA packaging. Finally, the plausible regulation of histone expression at the gene and protein levels in A. pacificum is discussed.

• Korean Journal of Microbiology
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• v.27 no.1
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• pp.21-26
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• 1989

The chromosomal DNA of Klebsiella penumoniae KNF1 was partially digested with HindIII. pBR322 was completely digested with same enzyme with additional alkaline phosphatase treatment. Both DNA samples were ligated and transformed into E. coli KO60. Single coloneis of the transformed cells were isolated on N0free agar media. These colonies were ampicillin-resistant and tetracycline-sensitive. When the plasmids in transformants were cured, nitrogen-fixing activities were lost. Therefore, these transformants harbored recombinant plasmid including nif genes of K. pneumoniae. Nitrogenase activity of transformant was higher than or the same as that of K. pneumoniae KNF1.

• Journal of Industrial Technology
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• v.29 no.B
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• pp.121-125
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• 2009

New cellulase genes, named as CelV2 and CelN1, respectively, were isolated from Erwinia carotovora ATCC15713 and expressed in E. coli. The CelV2 and CelN1 gene were PCR amplified with degenerated primers and PCR products were sequenced and expressed in E. coli. Two new cellulase genes showed 97% homologies with previously reported Erwinia cellulase genes. The recombinant cellulase were purified with Ni-NTA column chromatography and its enzymatic properties were characterized. The optimum temperature of two enzymes were about $50^{\circ}C$ degree and optimum pH were around pH7.0. The newly isolated celluase genes could be used for enhancing substrate range of alcohol-producing bacteria such as Zymomonas mobilis.

• Journal of Microbiology and Biotechnology
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• v.17 no.10
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• pp.1688-1694
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• 2007

A gene encoding the mannanase of Bacillus subtilis WL-3, which had been isolated from Korean soybean paste, was cloned into Escherichia coli and the nucleotide sequence of a 2.7-kb DNA fragment containing the mannanase gene was subsequently determined. The mannanase gene, designated manA, consisted of 1,080 nucleotides encoding a polypeptide of 360 amino acid residues. The deduced amino acid sequence was highly homologous to those of mannanases belonging to glycosyl hydrolase family 26. The manA gene was strongly expressed in B. subtilis 168 by cloning the gene downstream of a strong B. subtilis promoter of plasmid $pJ27{\Delta}88U$. In flask cultures, the production of mannanase by recombinant B. subtilis 168 reached maximum levels of 300 units/ml and 450 units/ml in LB medium and LB medium containing 0.3% locust bean gum, respectively. Based on the zymogram ofthe mannanase, it was found that the mannanase produced by recombinant B. subtilis could be maintained stably without proteolytic degradation during the culture time.

• Korean Journal of Microbiology
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• v.26 no.1
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• pp.32-36
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• 1988

The yeast gene HOM3 encodes aspartokinase, which catalyses the first step (aspartate to and from beta-aspartyl phosphate) of common pathway to threonine and methionine. The yeast HOM3 gene expression is known to be regulated by threonine and methionine specific control, and also by general control of amino acid biosynthesis. Isolation and characterization of the HOM3 gene are essential for the molecular genetic study on its regulation of expression. A recombinant plasmid pSC3 (15.5kb, vector YCp50) has been cloned into E. coli HB101 from yeast genomic library through their complementing activity of HOM3 mutation in a yeast recipient strain M34-24B. Organization of the plasmid was characterized by delineation of restriction cleavage sites in the insert fragment.

• International Journal of Industrial Entomology and Biomaterials
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• v.14 no.2
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• pp.69-74
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• 2007

As the genome of B. mori is available in GenBank and the EST database of B. mori is expanding, identification of novel genes of B. mori was conceivable by datamining techniques and bioinformatics tools. In this study, we used the in silico cloning method to get the 6-Phosphogluconolactonase (6PGL) gene of B. mori and analysed with bioinformatics tools. The result was confirmed by RT-PCR and prokaryotic expression. The 6PGL cDNA comtains a 702 bp ORF. The deduced protein has 233 amino acid residues, with the predicted molecular weight of 25946. 72 Da, isoelectric point of 5.41, and contains conserved NagB domains. This gene has been registered in GenBank under the accession number EF198104.

• Applied Biological Chemistry
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• v.31 no.3
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• pp.267-273
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• 1988

Transformed, hybrid strains of the yeast Saccharomyces capable of simultaneous secretion of both glucoamylase and ${\alpha}-amylase$ have been produced. These strains can carry out direct, one-step assimilation of starch with conversion efficiency greater than 93% during a 5 day growth period. One of the transformants converts 92.8% of available starch into reducing sugars in only 2 days. Glucoamylase secretion by these strains results from expression of one or more chromosomal STA genes derived from Saccharomyces diastaticus. The strains were transformed by a plasmid(pMS12) containing mouse salivary ${\alpha}-amylase$ cDNA in an expression vector containing yeast alcohol dehydrogenase promoter and a segment of yeast $2{\mu}$ plasmid. The major starch hydrolysis product produced by crude amylases found in culture broths is glucose, indicating that ${\alpha}-amylase$ and glucoamylase act cooperatively.

• YAKHAK HOEJI
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• v.35 no.1
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• pp.22-29
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• 1991

pMB4 is a 2.4-kilobase plasmid of Staphylococcus aureus TR-1 that confers inducible resistance to the macrolide-lincosamide-streptogramin B(MLS) antibiotics. By subcloning studies, it was found that the MLS resistance determinant was located at 1.0Kb fragment between Sau3AI and TaqI sites. DNA sequence of the MLS resistant determinant, named ermC-4 was determined, and found to be highly homologous with that of ermC. Because the leader peptide sequence of ermC-4 was identical with that of ermC, the expression of the resistance gene is thought to be controlled by posttranscriptional attenuation in S. aureus TR-1.