• 대한임상검사과학회지
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• 제38권2호
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• pp.111-116
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• 2006

One of the cardinal findings of the renal diseases is proteinuria, which appears in the early phase of kidney diseases and is very important in diagnosis, prognosis and decision making in the treatment process and results of the treatment. The study subjects were 126 patients who visited the nephrology department of Kangbuk Samsung Hospital. Serum was requested for urine protein electrophoresis. Total protein was measured with Bayer Advia 1650 (Biuret). Quantitation of each fraction was done by multiplying the percentage of each fraction by the total protein. Serum creatinine and BUN were also measured with Bayer Advia 1650 (Jaffe and Urease). Serum protein EP was done with REP(rapid electrophoresis) using Helena Kit reagents (REP Ultra SPE Kit, Ponceau S stain, Acetic acid, Methanol, EP Control). Concentrated urine was used for urine protein EP. The SPSS package was used for statistics analysis. Percentage and quantitation of the level of albumin in renal diseases were significantly lower than those in healthy controls. Total protein was correlated with albumin. In terms of proportion, ${\alpha}1$-globulin, ${\alpha}2$-globulin, ${\beta}$-globulin, and ${\gamma}$-globulin fractions were increased in the disease group. But, in the quantified level, ${\alpha}2$-globulin was increased and ${\beta}$-globulin and ${\gamma}$-globulin were decreased. ESRD patients showed an increased secretion of high molecular proteins in urine protein EP. A decreased level in serum total protein correlated with the decreased level of serum albumin and the total amount of urine total protein. This study revealed the variety in the level of serum and urine proteins and their subgroups by EP.

• Journal of Microbiology and Biotechnology
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• 제10권3호
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• pp.312-320
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• 2000

Abstract The full sequence of the plasmid pKJ36, which was derived from Bifidobacterium longum KJ, was determined and analyzed to construct shuttle vectors between E. coli and Bifidobacterium. The plasmid pKJ36 was composed of 3,625 base pairs with a 65.1% G+C content. The structural organization of pKJ36 was highly similar to that of pKJ50, and the three major ORFs on pKJ36 showed high amino acid sequence homologies with those of pKJ50. The putative proteins coded by these three ORFs were designated as RepB (32.0 kDa, pI=9.25), MembB (29.0 kDa, pI=12.25), and MobB (39.0 kDa, pI=IO.66), respectively. The amino acid sequence of RepB showed a 57% identity and 70% similarity with that of the RepA protein of pKJ50. Upstream of the repB gene, the so-called iteron sequence was directly repeated four-and-ahalf times and a conserved dnaA box was identified. An amino acid sequence comparison between the MobB and MobA of pKJ50 revealed a 48% identity and 61 % similarity. A conserved oriT sequence with an inverted repeat identical to that of pKJ50 was also found upstream of the mobB gene. A hydropathy analysis of MembB revealed four possible transmembrane regions. The expressions of the repB and membB genes were confirmed by RT-PCR. The in vitro translation reaction of pKJ36 showed protein bands with anticipated sizes with respect to each putative gene product. S 1 endonuclease treatment and Southern hybridization suggested that pKJ36 replicates by a rolling circle mechanism via a single-stranded DNA (ssDNA) intermediate. A shuttle vector between E. coli and Bifidobacterium sp. was constructed using the pKJ36, pBR322, and staphylococcal chloramphenicol acetyl transferase (CAT) gene. The successful transformation of the Bifidobacterium strains was shown by Southern hybridization and PCR. The transformation efficiency differed from strain to strain and, depending on the electroporation conditions, with a range between $1.2{\times}10^1-2.6{\times}10^2{\;}cfu/\mu\textrm{g}$ DNA.X> DNA.

• Journal of Microbiology and Biotechnology
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• 제9권1호
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• pp.91-97
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• 1999

In this work, a 1.9-kb region of enterococcal plasmid p703/5 was isolated and the nucleotide sequence analysis of the region was performed. One major open reading frame (ORF) was identified encoding a polypeptide of 28 kDa. Database comparisons suggested that the protein showed some homology with other bacterial RepA proteins. Upstream of the ORF, a potential dnaA box, AT-rich region and 22-bp tandemly repeated sequences (DNA iterons), a feature typical for many replication ori sites, were recognized. Deletion analysis using Exonuclease III and several restriction enzymes indicated that the three elements and the gene product from the ORF were essential for replication and that the minimum unit of DNA required for replication resided on the 1.2-kb AvaII subfragment. Thus, this gene product was referred to as RepA.

• 약학회지
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• 제43권6호
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• pp.789-792
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• 1999

The complete nucleotide sequence of pKH4, a small multidrug resistance (smr) plasmid isolated from multidrug resistant Staphylococcus aureus SA5, was determined. Sequence analysis has revealed that pKH4 has two open reading frames for Rep and Smr proteins. The comparison of the amino acid sequence of Smr protein of pKH4 with those of other Smr proteins of various Staphylococcus showed that Smr protein of pKH4 is a new member of the SMR family.

• 한국미생물·생명공학회지
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• 제26권6호
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• pp.566-568
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• 1998

Partial nucleotide sequence (nt 1-1842) of chloramphenicol resistance plasmid pKH7 has been reported previously and residual nucleotide sequence (nt 1843-4118) of pKH7 was determined and then the complete nucleotide sequence of pKH7 was obtained. pKH7 consists of 4118 bp and has three ORFs. Besides Rep and CAT proteins described in previous paper, Pre protein which mediates site-specific recombination in Staphylococcus aureus was found to be on pKH7. R $S_{A}$, a site-specific recombination site of Pre protein, and palA, a specific lagging-strand conversion signal, was also found in pKH7. Amino acid sequence of Pre protein of pKH7 was compared with those of other antibiotic resistant Staphylococcus aureus plasmids.s.

• Journal of Microbiology and Biotechnology
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• 제23권6호
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• pp.837-842
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• 2013

A cryptic plasmid, pMBLR00, from Leuconostoc mesenteroides subsp. mesenteroides KCTC 3733 was isolated, characterized, and used for the construction of a cloning vector to engineer Leuconostoc species. pMBLR00 is a rolling circle replication plasmid, containing 3,370 base pairs. Sequence analysis revealed that pMBLR00 has 3 open reading frames: Cop (copy number control protein), Rep (replication protein), and Mob (mobilization protein). pMBLR00 replicates by rolling circle replication, which was confirmed by the presence of a conserved double-stranded origin and single-stranded DNA intermediates. An Escherichia coli-Leuconostoc shuttle vector, pMBLR02, was constructed and was able to replicate in Leuconostoc citreum 95. pMBLR02 could be a useful genetic tool for metabolic engineering and the genetic study of Leuconostoc species.

• KSBB Journal
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• 제8권2호
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• pp.133-142
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• 1993

효모균에서의 유전자 재조합에 의한 단백절 생산에 미치는 유전학적, 환경적인 요인의 영향을 연구하였. Plasmid 안정도와 개수는 $REP^+$ 체계 에서 대단히 높은 반면, rep 체계에서는 매우 낮았다. $2{\mu}m$ circle plasmid genome을 포함하는 plasmid의 경우에 있어서. $[cir^o]$ 형 세포에서의 plasmid 안정도와 개수가 $[cir^+]$형 세포에서보다 높기때문에 $[cir^+o]$형 세포가 더 선호되는 세포였다. 유전자 발현은 plasmid 개수와 안정도에 좌우 되었다. 촉진제의 양이 유전자 발현에 매우 중요 한 역할을 했다. 유전자 발현의 촉진에 필요한 g떠actose의 농도는 0.8% 이 변 충분했다. 높은 안정 도와 개수를 갖는 plasmid의 경우 촉진속도는 매우 빨랐다. Galactose가 배양의 시작부분부터 첨가 될 때가 mid-exponential ph잃e에 첨가될 때보다 유전자 발현의 극대점에 이르는 시간이 걸었다. 상대적 촉진제의 양이 증가함에 따라 glucc잉e억제 현상은 감소되었다.

• Journal of Microbiology and Biotechnology
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• 제21권8호
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• pp.796-801
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• 2011

Corynebacterium tuberculostearicum B146, a strain derived from healthy human skin, contains a medium copy plasmid, p1B146. This plasmid was cloned and its complete nucleotide sequence determined. As a result, p1B146 was found to be 4,2 kb in size with a 53% G+C content, plus six open reading frames (ORFs) were distinguished. According to a computer-assisted alignment, two of the ORFs exhibited significant similarities to already-known common plasmid proteins, the first being the RepA gene, responsible for plasmid replication via a rolling-circle mechanism, and the second being an FtsK-like protein, the function of which remains unclear. The presence and quantity of RNA fragments in the putative ORFs were also evaluated.

• 미생물학회지
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• 제52권1호
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• pp.110-115
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• 2016

남극 해양세균 Pseudoalteromonas sp. PAMC 21150에서 분리한 소형 플라스미드(small plasmid, pDK4)의 크기는 3,480bp이고 G+C 함량은 41.64%이며, 3개의 open reading frames(ORFs)을 포함하고 있다. 3개의 ORF는 replication initiation protein (RepA), conjugative mobilization protein (Mob), 그리고 기능이 밝혀지지 않은 단백질을 코팅하고 있다. PCR 반응으로 증폭한 pDK4를 Escherichia coli high-copy pUC19 클로닝 벡터에 삽입하여 fusion vector (pDOC153)를 제작하였고, pDOC 153에 chloramphenicol 저항성 유전자를 삽입하여 ampicillin/chloramphenicol 저항성 Pseudoalteromonas - Escherichia coli 셔틀 벡터(shuttle vector; 7,216 bp 크기; pDOC155)를 제작하였다. 북극 해양세균 P. issachenkonii PAMC 22718이 보유한 2개의 유전자(TonB-dependent receptor gene, chi22718_IV, and exochitinase gene, chi22718_III)를 pDOC155에 삽입하여 두 개의 pDOC155 변형체(pDOC158, pDOC165)를 제작하였다. pDOC158 혹은 pDOC165을 이용하여 triparental mating 방법에 의해 플리스미드 미보유 해양세균인 Pseudoalteromonas sp. PAMC 22137를 형질전환하였다. PCR을 이용한 유전자 증폭실험을 통해서, pDOC158와 pDOC165에 삽입된 유전자들은 Pseudoalteromonas sp. PAMC 22137와 E. coli $DH5{\alpha}$ 내에서 안정적으로 유지되는 것을 확인하였다. 위의 결과는 셔틀 벡터 pDOC155는 Pseudoalteromonas spp. 유래 유전자들을 다른 Pseudoalteromonas spp. 세포 안으로 전달할 수 있는 새로운 유전자 전달시스템으로 이용될 수 있음을 보여주었다.

• Food Science and Biotechnology
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• 제18권2호
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• pp.396-401
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• 2009

The nucleotide sequence contains 2 open reading frames encoding a 45-amino-acid protein homologous to a transcriptional repressor protein CopG, and a 203-amino-acid protein homologous to a replication protein RepB. Putative countertranscribed RNA, a double-strand origin, and a single-strand origin were also identified. A shuttle vector, pUCL2.1, for various lactic acid bacteria (LAB) was constructed on the basis of the pCL2.1 replicon, into which an erythromycin-resistance gene as a marker and Escherichia coli ColE1 replication origin were inserted. pUCL2.1 was introduced into E. coli, Lc. lactis, Lactobacillus (Lb.) plantarum, Lb. paraplantarum, and Leuconostoc mesenteroides. The recombinant LAB maintained traits of transformed plasmid in the absence of selection pressure over 40 generations. Therefore, pUCL2.1 could be used as an E. coli/LAB shuttle vector, which is an essential to engineer recombinant LAB strains that are useful for food fermentations.