대장균의 형질전환을 위한 Electroporation 最適條件

서승염,김준기,김주연 公州大學校 基礎科學硏究所 1995 自然科學硏究 Vol.4 No.-

형질전환을 위한 기존의 electroporator들이 일회용 cell과 고전압을 사용하지만, 한국생공에서 만들어진 Jetgene electroporator는 영구용 cell과 저전압을 사용하여 형질전환할 수 있도록 한 한국실정에 맞는 새로운 기기이다. 이 기기를 사용하여 대장균의 형질전환을 위한 electroporation 최적조건을 찾기 위하여 전기장의 세기, 펄스의 길이, plasmid DNA의 크기, 균주, DNA 농도, 세포의 농도, 세척 용액과 횟수, 배지의 종류가 Electroporation의 효율에 영향을 미치는 영향을 조사했다. 전기장의 세기는 12.14 - 12.85 kV/cm에서, 시정수 (τ)가 4-5 msec에서, 조사한 대장균주 중에서 BW 13745에서 최고의 효율을 나타냈다. DNA 양에 형질전환 효율이 비례적으로 증가하지는 않았지만 DNA 양이 많아질수록 많은 형질전환체가 얻어져 DNA 1-2㎍에서 109의 형질전환체가 얻어졌다. 수확시 세포의 밀도가 OD600이 0.6-0.8일 때 최고의 효율을 나타냈다 여러 세척액 중에서 10 % glycerol로 세척했을때, 2회 세척했을 때, 여러 배지 가운데 SOC에서 자란 균을 사용했을 때 최고의 효율을 나타냈다. 이 조건들을 조합해 실험을 해본 결과 DNA 1㎍당 1점에서 1010까지의 형질전환체를 얻을 수 있었다. 다른 electroporator에 비하면 상당히 저가인 한국생공의 Jetgene electroporator를 사용하여 적은 비용으로 높은 효율의 형질전환을 할 수 있었다. Although disposable cells and high voltage are used for electroporators, a new electroporator for which permanent cells and low voltage can be used was manufactured from Korea Biotech. Inc We wanted optimize the procedures for electroporation using this electroporator. The effect of electric field strength, pulse length, sizes of plasmid, host strains, washing solutions and times, growth media were studied. Maximum transformation efficiency for E. coli strain CC118 was obtained at electric field strength of 12.14-12.85 kv/cm and retention time of 4-5 msec. Although the number of transformants was not proportional to the amount of plasmid DNA, larger amount of DNA gave higher number of transformants. 109 transformants were obtained with 1-2㎍ DNA. The highest number of transformants were obtained using cells harvested at OD600 0.6-0.8. 10% glycerol out of washing solutions, 2 times washing, SOC medium out of growth media were best for high efficiency of transformation. 109-1010 transformants were routinely obtained. Jetgene electroporator seems adequate for electroporation at lower price and working cost.

Evidence for two $Na^+$/$H^+$ Antiport Systems in Escherichia coli

Seo, Sung-Yum The Microbiological Society of Korea 1992 미생물학회지 Vol.30 No.4

Several insertion mutants of Escherichia coli in the ant gene, coding for $Na^{+}$ $H^{+}$ antiport activity, showed littel, if any, reduction in the antiport activity. $Na^{+}$ dependent transport activity also remained at wild type level. These facts led to the idea that E. coli has evolved at least two distinct systems for extrusion of $Na^{+}$ The antiport activities were studied under various conditions to reveal different properties of these systems. For convenience these activities are referred to as major and minor activities. The distinguishing properties of the two systems include : kinetics (Km, Vm) at pH 7.8, competition pattern between $Na^{+}$ and Li$^{+}$ , pH profiles, pattern of the change in kinetic parameters as a function of pH, and sensitivity to protease, chemicals and heat.

Genetic and Biochemical Study of the ant Gene , Coding a Specialized Na+ / H+ Antiport System of Escherichia coli

SUNG YUM SEO 생화학분자생물학회 1992 생화학분자생물학회 춘계학술발표논문집 Vol.- No.-

Regulation of the expression of nhaA gene coding $Na^{+}$/$H^{+}$ antiporter A of escherichia coli

Seo, Sung-Yum,Lee, Seung-Heon The Microbiological Society of Korea 1995 The journal of microbiology Vol.33 No.2

.betha.-galactosidase activity of Escherichia coli cells containing operon fusion nhaA'-'lacZ was monitored to study the regulation of expression of nhaA gene under various conditions. The expression of the fusion was enhanced only by chemicals containing Na$^{+}$ or Li$^{+}$. This Na$^{+}$ or Li$^{+}$. This Na$^{+}$(Li$^{+}$)-specific enhancement of .betha.-galactosidase activity represented the increase in the rate of synthesis of .betha.-galactosidase rather than the decrease in the breakdown rate. The induction pattern was influenced by copy numbers of the gene. Induction by Na$^{+}$ or Li$^{+}$ was concentration and time dependent, reaching maximum 5-6 fold induction after 2 hours at 0.4-0.5 M for Na$^{+}$ or at 0.25-0.35 M for Li$^{+}$, Although the expression was induced at much lower concentration of Na$^{+}$ at alkaline pH values than at neutral pH in the presence of Na$^{+}$, alkaline pH itself did ot induced the expression of the fusion in the absence of Na$^{+}$. Temperature shift and growth phase of culture did not affect the level of induction.he level of induction.

대립계 포도 천창개폐형 비가림하우스의 최적 파이프 규격

염성현(Sung Hyun Yum),윤남구(Nam Gyu Yun),김경원(Gyeong Won Kim),이성현(Sung Hyoun Lee),조용호(Yong Ho Cho),박서준(Seo Jun Park),박문균(Mun Kyun Park) (사)한국생물환경조절학회 2007 생물환경조절학회지 Vol.16 No.4

본 연구는 기존 비닐하우스 아연도 강관을 사용한 하우스 폭 3.6m와 5m 천창개폐형 대립계 포도 비가림하우스에 대한 구조적 안전성을 검토하고, 인장강도 400Nㆍ㎜?²(SGH400 등) 이상의 파이프를 사용하는 조건에서 하우스 폭 5m인 천창개폐형 대립계 포도 비가림하우스에 대하여 구조적으로 안전한 최적 파이프 규격을 제시하고자 수행하였다. 주기둥 3m×서까래 60 ㎝인 천창개폐형 3.6m 비가림하우스의 경우, 적설심 35㎝에서는 구조적으로 안전한 것으로 분석되었으나 측면 및 전후면 풍속 35mㆍs?¹에서는 불안전한 것으로 나타났으며, 동일 주기둥과 서까래 간격을 갖는 천창개페형 5m 비가림하우스의 경우에는 적설심 35와 풍속 35mㆍs?¹에서 모두 불안전하여 구조보강이 필요한 것으로 분석되었다. 그리고 동일 주기둥과 서까래 간격을 가지나 인장강도 400Nㆍ㎜?² 이상을 갖는 파이프를 사용하는 조건에서 천창개폐형 5m 비가림하우스의 최적 파이프 규격은 지붕높이 1.6m(아치형)와 지붕높이 1.8(복숭아형)에 대하여 동일하게 두 경우로 규격화할 수 있었다. 즉, 안전풍속 35mㆍs?¹와 안전적 설심 40㎝에서 구조적으로 안전한 서까래 규격은 Ф31.8×1.5t@600이었으며, 안전풍속 30mㆍs?¹와 ss 안전적설심 35㎝에서는 서까래 Ф25.4×1.5t@600인 것으로 분석되었다. 덕면으로부터 곡부보까지의 높이는 안전적설심보다는 안전풍속에 직접적인 영향을 미치는 것으로 분석되었으며, 처마를 높임에 따라 측면풍속에 대해서는 방풍벽파이프(측벽서까래)를, 전후면 풍속에 대해서는 마구리기둥의 규격을 강화하여야 하는 것으로 분석되었다. This study was carried out to: (1) analyze structural stability of representative rainsheltering greenhouses for large-grain grapevine cultivation with widths of 3.6 and 5 m in case of using the existing pipe for agriculture; (2) present the optimum specification of pipes in the greenhouse with a width of 5 m under the condition of using the pipe of which ultimate strength has been above 400 Nㆍ㎜?²; (3) evaluate stability and also present the optimum specification of pipes as eaves height was augmented. The above analyses were done for greenhouses with roof vents and also with a main-column interval of 3 m and a rafter interval of 60㎝ First, the existing 3.6m greenhouse with a rafter of Ф25.4 × 1.5 t@600 was stable for a snow-depth of 35 ㎝ but unstable for a wind velocity of 35mㆍs?¹, Meanwhile the existing 5 m greenhouse with the same rafter was not stable for a wind velocity of 335 mㆍs?¹ as well as a snow-depth of 35㎝. This meant that existing greenhouses had to be reinforced to secure stability. Second, the specification of pipes, especially rafter, could be classified as two cases. One had a structural stability at a safe wind velocity of 35 mㆍs?¹ and a safe snow-depth of 40㎝ for which stability the rafter had to be Ф31.8 × 1.5 t@600, and the other had a stability at 30 mㆍs?¹-35㎝ at the specification of rafter Ф25.4 × 1.5 t@600. Finally, eaves height had a significant effect on safe wind velocity. But it had little influence on safe snow-depth. The results showed that the specification of side-wall pipes had to be reinforced for the safe side velocity according to the increment of eaves height and similarly the specification of fore?end post for the safe fore-end velocity.

대립계 포도 천창개폐형 비가림하우스의 최적 파이프 규격

염성현(Sung Hyun Yum),윤남규(Nam Gyu Yun),김경원(Gyeong Won Kim),이성현(Sung Hyoun Lee),조용호(Yong Ho Cho),박서준(Seo Jun Park),박문균(Mun Kyun Park) (사)한국생물환경조절학회 2005 생물환경조절학회지 Vol.14 No.2

본 연구는 기존 비닐하우스 아연도 강관을 사용한 하우스 폭 3.6m와 5m 천창개폐형 대립계 포도 비가림히우스에 대한 구조적 안전성을 검토하고, 인장강도 400Nㆍ㎜?²(SGH400 등) 이상의 파이프를 사용하는 조건에서 하우스 폭 5m인 천창개폐형 대립계 포도 비가림하우스에 대하여 구조적으로 안전한 최적 파이프 규격을 제시하고자 수행하였다. 주기둥 3m×서까래 60㎝인 천창개폐형 3.6m 비가림하우스의 경우, 적설심 35㎝에서는 구조적으로 안전한 것으로 분석되었으나 측면 및 전후면 풍속 35mㆍs?¹에서는 불안전한 것으로 나타났으며, 동일 주기둥과 서까래 간격을 갖는 천창개폐형 5m 비가림하우스의 경우에는 적설심 35와 풍속 35mㆍs?¹에서 모두 불안전하여 구조보강이 필요한 것으로 분석되었다. 그리고 동일 주기둥과 서까래 간격을 가지나 인장강도 400Nㆍ㎜?² 이상을 갖는 파이프를 사용하는 조건에서 천창개폐형 5m 비가림하우스의 최적 파이프 규격은 지붕높이 1.6m(아치형)와 지붕높이 1.8m(복숭아형)에 대하여 동일하게 두 경우로 규격화할 수 있었다. 즉, 안전풍속 35mㆍs?¹와 안전적설심 40㎝에서 구조적으로 안전한 서까래 규격은 φ31.8 × 1.5t@600이었으며, 안전풍속 30mㆍs?¹와ss 안전 적설심 35㎝에서는 서까래 φ25.4 × 1.5t@600인 것으로 분석되었다. 덕면으로부터 곡부보까지의 높이는 안전적설심보다는 안전풍속에 직접적인 영향을 미치는 것으로 분석되었으며, 처마를 높임에 따라 측면풍속에 대해서는 방풍벽파이프(측벽서까래)를, 전후면 풍속에 대해서는 마구리기둥의 규격을 강화하여야 하는 것으로 분석되었다. This study was carried out to: (1) analyze structural stability of representative rain-sheltering greenhouses for large-grain grapevine cultivation with widths of 3.6 m and 5 m in case of using the existing pipe for agriculture; (2) present the optimum specification of pipes in the greenhouse with a width of 5 m under the condition of using the pipe of which ultimate strength has been above 400 Nㆍ㎜?²; (3) evaluate stability and also present the optimum specification of pipes as eaves height was augmented. The above analyses were done for greenhouses with roof vents and also with a main-column interval of 3 m and a rafter interval of 60 ㎝, First, the existing 3.6 m greenhouse with a rafter of φ25.4 × 1.5 t@600 was stable for a snow-depth of 35 ㎝ but unstable for a wind velocity of 35 mㆍs?¹. Meanwhile the existing 5 m greenhouse with the same rafter was not stable for a wind velocity of 335 mㆍs?¹ as well as a snow-depth of 35 ㎝. This meant that existing greenhouses had to be reinforced to secure stability. Second, the specification of pipes, especially rafter, could be classified as two cases. One had a structural stability at a safe wind velocity of 35 mㆍs?¹ and a safe snow-depth of 40 ㎝ for which stability the rafter had to be φ31.8 × 1.5 t@600, and the other had a stability at 30 mㆍs?¹-35 ㎝ at the specification of rafter φ25.4 × 1.5 t@600. Finally, eaves height had a significant effect on safe wind velocity. But it had little influence on safe snow-depth. The results showed that the specification of side-wall pipes had to be reinforced for the safe side velocity according to the increment of eaves height and similarly the specification of fore-end post for the safe fore-end velocity.

Escherichia coli 의 Na+/H+ antiporter 유전자(ant)에 있어서 sodium 과 alkalinity 감수성 돌연변이체 mapping 의 분리 및 특성

서승염 한국유전학회 1992 Genes & Genomics Vol.14 No.2

Mutants of Escherichia coli displaying a pleiotropic phenotype including sensitivity to Na^+, Li^+, and alkaline pH were isolated using insertion mutagenesis (Tn10 and λplacMU53). Genetic mapping by both conjugation (F and Hfr) and transduction (Pl) located the mutations near the region of the Na^+/H^+ antiporter gene(s). The mutations were further pinpointed via complementation using recombinant plasmid pNHA7. The presence of the ant gene alone on this plasmid is sufficient for complementation. To study whether the ant gene is a regulatory of structural gene, in vitro mutagenesis was used to generate a class of mutants displaying altered transport of Na^+ versus Li^+. These results suggest that E. coli has evolved two or more Na^+/H^+ antiporter genes including ant as well as other unmapped systems.

Replacement of Chromosomal nha4 Gene of Escherichia coli with in Vitro Mutated Gene

서승염 한국유전학회 1995 Genes & Genomics Vol.17 No.2

Designed deletion and insertion were constructed on plasmids into nhaA gene, coding a Na^+/H^+ antiporter A (NhaA) of Escherichia coli. The chromosomal nhaA gene was replaced with the mutated genes, resulting deletion and insertion mutants. The designed mutations in chromosomal nhaA genes of the mutants were confirmed by genetic analysis and after retrieval of the chromosomal mutations on plasmids. The deletion and insertion mutants displayed sodium (lithium) and alka-linity sensitive phenotypes. The Na^+/H^+ antiport activity of the mutants is not significantly altered, due to another Na^+/N^+ antiporter (NhaB) in E. coli (Seo, 1992, Kor. J. Microbiol. 30 : 269-277).

Low infectivity of a novel avian-origin H7N9 influenza virus in pigs

Yum, Jung,Park, Eun Hye,Ku, Keun Bon,Kim, Ji An,Oh, Seung Kyoo,Kim, Hyun Soo,Seo, Sang Heui Springer-Verlag 2014 Archives of virology Vol.159 No.10

수술 전 심전도분석과 어린이 사시 수술 중 서맥의 발생정도와의 상관관계

서광석,김종성,김희수,정용보,염명걸 대한마취과학회 1999 Korean Journal of Anesthesiology Vol.37 No.5

Background : Severe bradycardia occurring in strabismus surgery is unpredictable and may lead to cardiac arrest. If we identify patients who have risk of severe bradycardia during strabismus surgery, it is helpful to provide a more meticulous anesthesia. In this study, we investigated the correlation between the degree of bradycardia and indices of power spectral analysis and nonlinear dynamic data analysis of preoperative ECG. Methods : ECG was collected for twenty minutes from strabismus patients (n = 93). No premedicants were administered. After administration of anesthesia and traction of extraocular muscle, the lowest heart rate was recorded. We calculated the correlation coefficient between the power spectral density, approximate entropy and correlation dimension of preoperative ECG and the difference between preoperative and lowest HR. Results : As the difference between the preoperative heart rate and the intraoperative bradycardia increased, the preoperative ECG showed a low total power (r = 0.27, P = 0.016), low low-frequency power (r = 0.31, P = 0.049), low high-frequency power (r = 0.30, P = 0.007) and high ratio of low to high-frequency power (r = 28, P = 0.03). There was no correlation between bradycardia and approximate entropy and correlation dimension, respectively. Conclusions: There was a correlation between indices of power spectral analysis of preoperative ECG and degree of bradycardia during strabismus surgery. Therefore we take into consideration preoperative ECG and its analytic indices in order to provide careful preparation for high risk patients who exhibit a tendency to severe bradycardia. (Korean J Anesthesiol 1999; 37: 739∼744)