• 제목/요약/키워드: iClone

검색결과 204건 처리시간 0.027초

Cloning and Nucleotide Sequence of a cDNA Encoding the Rat Triosephosphate Isomerase

  • Lee, Kyunglim;Ryu, Jiwon
    • Archives of Pharmacal Research
    • /
    • 제19권6호
    • /
    • pp.497-501
    • /
    • 1996
  • A gene coding for triosephosphate isomerase (TPI) from a rat skeletal muscle cDNA library was cloned and its nucleotide sequence was determined. The 1, 348-bp cDNA clone contains 24 bp $5^I$ noncoding region, the entire 750 bp coding region corresponding to a protein of 249 amino acids, $547bp 3^I$ noncoding region and part of a poly(A) tail. It also contains a polyadenylation signal, AATAAA, starting from 17 bp upstream of the poly(A) tail. The calculated molecular weight of rat TPI is 27.8 kDa and the net charge is +4. The deduced amino acid sequence from rat TPI CDNA sequence has 93% and 94% homology with that of mouse and human clones, respectively. The amino acids at the residue of Asn12, Lys14, His96, Glu 166, His96, His101, Ala177, Tyr165, Glu13O, Tyr2O9, and Ser212 in catalytic site are completely identical, confirming that the functional residues in TPI proteins are highly conserved throughout evolution. The most profound characteristic of rat TPI enzyme, compared with other TPIs, is that there are five cysteine substitutions at the residue of 21, 27, 159, 195 and 204. A Glu123 instead of Gly was found in rabbit, rhesus, mouse and human sequences. Through the method of RT-PCR, the mRNA transcription level of TPI gene was found to be different among various tissues and was highest in muscle.

  • PDF

Actinodura roseorufa에서 생산되는 UK-58,852로부터 PKS type I 에 관련된 생합성 유전자의 분리 및 분석

  • 김자용;이주호;김대희;김동현;송재경;이희찬
    • 한국생물공학회:학술대회논문집
    • /
    • 한국생물공학회 2000년도 추계학술발표대회 및 bio-venture fair
    • /
    • pp.660-664
    • /
    • 2000
  • UK-58,852의 생합성에 관여하는 유전자를 분리하기 위해 Actinomadura roseorufa의 genomic DNA와 E. coli-Streptomyces shuttle cosmid vector인 pOJ446이 genomic library를 만들었다. Genomic library는 dehydratase PCR product와 eryA 유전자를 probe로 하여 sugar 생합성 유전자와 polyketide typel 유전자가 집단으로 존재하는 cosmid pHD54를 분리하였고, 이를 제한 효소인 BamHI, SmaI와 Sonicater를 이용해서 subcloning 하였다. 이들의 염기서열을 부분 분석한 결과, polyketide 생합성에 관여하는 ketoacyl synthase, methylmalonyl acyltransferase, ketoreductase, enolreductase 그리고 PKS loading domain 등 polyketide synthase type I 임을 보여주고 있고, BLAST 분석된 결과를 보면 polyketide synthase 유전자는 rifamycin 생합성 유전자와 유사성이 높다. 그리고 sugar 생합성에 관여하는 유전자로는 oxidoreductase, dTDP-D-glucose 4,6 dehydratase, dTDP-D-glucose synthase 그리고 dTDP-4-keto-6-deoxy-D-glycose 3,5-epimerase으로 구성된 gene cluster를 확인하였다. 그리고 염기서열 분석된 유전자중 dTDP-D-glucose synthase를 발현하여 유전자의 기능을 확인하였다.

  • PDF

배추로부터 광계 I의 PSI-H Subunit Homolog의 클로닝 및 분자생물학적 특성 연구 (Molecular Cloning and Characterization of a cDNA for the PSI-H Subunit Homolog of Photosystem I in Chinese Cabbage)

  • 차준영;최영진;이효신;김기용;박근제;조진기;손대영
    • 한국초지조사료학회지
    • /
    • 제22권1호
    • /
    • pp.51-58
    • /
    • 2002
  • 식물의 광합성에 관여하는 광계 I의 protein subunit들의 연구는 최근까지도 극히 미약한 실정이며, 각각의 subunit들의 특성 또한 일부만이 밝혀져 있다. 본 연구진은 배추의 cDNA library로부터 식물에만 존재하는 subunit 중의 하나인, PSI-H subunit을 암호화하는 유전자인 bpsaH를 분리하였다. 이 유전자는 총 633 bp의 염기로 구성되어 있으며, 염기서열로부터 추정되는 분자량은 약 15,400이었고 등전점은 9.91이었다. 배추 PSI-H subunit의 아미노산 서열을 다른 식물체 유래의 단백질들과 비교분석한 결과, 시금치의 PSI-H와 가장 높은 유사성 (79.3%)을 나타내었다. 또한 bpsaH의 조직 특이적 발현 양상을 조사한 결과, 광합성 조직인 잎에서는 강하게 발현된 반면 꽃봉우리에서는 약하게 발현되었으며, 비광합성 조직인 뿌리에서는 전혀 발현되지 않았다.

질소시용, 예초 및 재식밀도가 한국잔디(Zoysia Japonica Steud)의 생육에 미치는 영향 (Studies on the growth of Korea Lawn Grass (Zoysia japonica Steud.)in Reponse to Nitrogen Application, Clipping Treatment and Plant Density)

  • 심재성
    • 자연과학논문집
    • /
    • 제1권
    • /
    • pp.61-113
    • /
    • 1987
  • The increasing emphasis placed on the production of fine turf for lawns, golf courses, parks, and other recreational sites has led to many unsolved problems as to how such turf could be best established and mainteined. For this purpose, a series of experiments were conducted under con ditions of pot and field. The results obtained were as follows EXPERIMENT I. The effect of nitrogen fertilizer and clipping interval on Zoysia japonica. 1. Increasing the rate of nitrogen and frequent clipping increased tiller number of Zoysis japonica and the maximum number of tillers were obtained from 700 kg N application and freqnent clippings (10 days interval ) in October. Treatment of 350kg N with 10 days clipping interval increased tillers much more than those of 700 kgN with 20 and 30 days clipping intervals. 2. The average number of green leaves occurred during the growth period maximized by applying 700 kg N and clipping 10 days interval. 3. Increasing tiller numbers significantly decreased tops DM weight per tiller by clipping plants at interval of 10 and 20 days, irrespective of nitrogen applied, and with nil N, at the interval of 30 days. By applying 700 kg N, however, top DM weight per tiller increased as the number of tillers increased consistently. 4. The highest top DM weight was achieved from late August to early September by applying 350 and 700kgN. 5. During the growth period, differences in unders ( stolon + root ) DM weight occurred bynitrogen application were found between nil N and two applied nitrogen levels, whereas, at the same level of nitrogen applied, the increase in stolon DM weight enhanced by lengthening the clipping interval to 30 days. 6. Nitrogen efficiency to green leaves, stolon nodes and DM weight of root with high nitrogen was achieved as clipping interval was shortened. 7. By increasing fertilizer nitrogen rate applied, N content n the leaves and stems of Zoysiajaponica was increased. On the other hand, N content in root and stolon had little effect onfertilizer nitrogen, resulting in the lowest content among plant fractions. The largest content of N was recorded in leaves. Lengthening the clipping interval from 10 or 20 to 30 days tends to decrease the N content in the leaves and stems, whereas this trend did not appeared in stolon androot. 8. A positive correlations between N and K contents in tops and stolon were established andthus K content increased as N content in tops and stolon increased. Meanwhile, P content was not affected by N and clipping treatments. 9. Total soluble carbohydrate content in Zoysia japonica was largest in stolon and stem, and was reduced by increasing fertilizer nitrogen rate. Reduction in total soluble carbohydrate due to increased nitrogen rate was severer in the stolons and stems than in the leaves. 10. Increasing the rate of nitrogen applied increased the number of small and large vascular bundles in leaf blade, but shortened distance among the large vascular bundles. Shortening the clipping interval resulted in increase of the number of large vascular bundles but decrease ofdistance between large vascular bundles.EXPERIMENT II. Growth response of Zoysia japonica imposed by different plant densities. 1. Tiller numbers per unit area increased as plant density heightened. Differences in num ber between densities at higher densities than 120 D were of no significance. 2. Tiller numbers per clone attained by 110 days after transplanting were 126 at 40D,77 at 80D, 67 at 120D, 54 at 160D, and 41 at 200D. A decreasing trend of tiller numbers per clone with increasing density was noticable from 100 days after transplanting onwards. 3. During the growth period, the greatest number of green leaves per unit area were attainedin 90days after transplanting at 160D and 200D, and 100 days after transplanting at 40D, 80Dand 120D. Thus the period to reach the maximum green leaf number with the high plantdensity was likely to be earlier that with the low plant density. 4. Stolon growth up to 80 days after transplaning was relatively slow, but from 80 daysonwards, the growth quickened to range from 1.9 m/clone at 40D to 0.6m/clone at 200Din 200 days after transplanting, these followed by the stolon node produced. 5. Plant density did not affect stolon weight/clone and root weight/clone until 80 daysafter transplanting. 6. DM weight of root was heavier in the early period of growth than that of stolon, butthis trend was reversed in the late period of growth : DM weight of stolon was much higherthan that of root.EXPERIMENT Ill. Vegetative growth of Zoysia japonica and Zoysia matrella as affected by nitrogen and clipping height. 1. When no nitrogen was applied to Zoysia japonica, leaf blade which appeared during theAugust-early September period remained green for a perid of about 10 weeks and even leavesemerged in rate September lived for 42 days. However, leaf longevity did not exceed 8 weeks asnitrogen was applied. In contrast the leaf longevity of Zoysia matrella which emerged during the mid August-earlySeptember period was 11 weeks and, under the nitrogen applied, 9 weeks, indicating that thelife-spen of individual leaf of Zoysia matrella may be longer than that of Zoysia japorica. Clipping height had no effect on the leaf longevity in both grasses. 2. During the July-August period, tiller number, green leaf number and DM weightof Zoysia japonica were increased significantly with fertilizer nitrogen, but were not with twolevel of clipping height. This trend was reversed after late September ; no effect of nitrogen wasappeared. Instead, lax clipping increased tiller number, green leaf number and DM weight. Greenleaves stimulated by lax clipping resulted in the occurrance of more dead leaves in late October. 3. Among the stolons outgrown until early September, the primary stolon was not influencedby nitrogen and clipping treatments to produce only 2-3 stolons. However, 1st branch stoIon asaffected by nitrogen increased significantly, so most of stolons which occurred consisted of 1st branch stolons. 4. Until early September, stolon length obtained at nil nitrogen level was chiefly caused bythe primary stolons. By applying nitrogen, the primary stolons of Zoysia japonica waslonger than 1st branch stolons when severe clipping was involved and in turn, shorter than 1stbranch stolons when lax clipping was concerned. In Zoysia matrella, 1st branch stolons were muchlonger than the primary stolon when turf was clipped severely but in conditions of lax clippingthere was little difference in length between primary and 1st branch stolons. 5. Stolon nodes of both Zoysia japonica and Z. matrella were positively influenced by nit rogen, but no particular increase by imposing clipping height treatment was marked in Zoysiamatrella. Although the stolon of Zoysia japonica grew until late October, the growthstimulated by nitrogen was not so remarkable as to exceed that by nil N.

  • PDF

Outbreaks of Imipenem-Resistant Acinetobacter baumannii Producing Carbapenemases in Korea

  • Jeong Seok-Hoon;Bae Il-Kwon;Park Kwang-Ok;An Young-Jun;Sohn Seung-Ghyu;Jang Seon-Ju;Sung Kwang-Hoon;Yang Ki-Suk;Lee Kyung-Won;Young Dong-Eun;Lee Sang-Hee
    • Journal of Microbiology
    • /
    • 제44권4호
    • /
    • pp.423-431
    • /
    • 2006
  • Among 53 Acinetobacter baumannii isolates collected in 2004, nine imipenem-resistant isolates were obtained from clinical specimens taken from patients hospitalized in Busan, Korea. Nine carbapenemase-producing isolates were further investigated in order to determine the mechanisms underlying resistance. These isolates were then analyzed via antibiotic susceptibility testing, microbiological tests of carbapenemase activity, pI determination, transconjugation test, enterobacterial repetitive consensus (ERIC)-PCR, and DNA sequencing. One outbreak involved seven cases of infection by A. baumannii producing OXA-23 ${\beta}-lactamase$, and was found to have been caused by a single ERIC-PCR clone. During the study period, the other outbreak involved two cases of infection by A. baumannii producing IMP-1 ${\beta}-lactamase$. The two clones, one from each of the outbreaks, were characterized via a modified cloverleaf synergy test and an EDTA-disk synergy test. The isoelectric focusing of the crude bacterial extracts detected nitrocefin-positive bands with pI values of 6.65 (OXA-23) and 9.0 (IMP-1). The PCR amplification and characterization of the amplicons via direct sequencing showed that the clonal isolates harbored $bla_{IMP-1}$ or $bla_{oxA-23}$ determinants. The two clones were characterized by a multidrug resistance phenotype that remained unaltered throughout the outbreak. This resistance encompassed penicillins, extended-spectrum cephalosporins, carbapenems, monobactams, and aminoglycosides. These results appear to show that the imipenem resistance observed among nine Korean A. baumannii isolates could be attributed to the spread of an IMP-lor OXA-23-producing clone. Our microbiological test of carbapenemase activity is a simple method for the screening of clinical isolates producing class D carbapenemase and/or class B $metallo-{\beta}-lactamase$, in order both to determine their clinical impact and to prevent further spread.

Bacillus thuringiensis serovar. darmstadiensis의 곤충치사독소 유전자분리 및 구조해석 (Isolation and Analysis of Bacillus thuringiensis serovar. darmstadiensis Insecticidal Protein Gene)

  • 김도영;구본성;도대홍
    • 한국식품영양학회지
    • /
    • 제9권4호
    • /
    • pp.459-465
    • /
    • 1996
  • 지금까지 많은 연구가 되어 있지 않던 Bacillus thuringiensis serovar. darmstadiensis의 내독소를 Renografin-76 단계적 기울기 원심분리로 분리하여 전자 현미경으로 관찰하여 이중피라미드 구조를 가진 독소 단백질을 확인하였으며 B. thuringiensis serovar. kurstaki HD1의 독소 생성유전자와 B. thuringiensis serovar. darmstadiensis의 유전자가 유사성이 있다는 보고를 근거로 하여 B. thuringiensis serovar. HD1의 독소 생성유전자를 가진 프로브(pUYBT 9044)로 이용하여 colony hybridization 및 Southern hybridization 한 결과 2.6Kb EcoRI 단편 및 3.6Kb Hind III 단편을 선발할 수 있었다. 이들 단편들은 B. thuringiensis serovar. kurstaki HD1 독소 유전자와 hybridization시 유사성이 있었다. 특히 3.5Kb HindIII 단편은 2.6Kb EcoR I 단편에 클로닝되어 있는 1.8Kb의 HD1 독소 유전자와 유사성이 있는 부분을 공유하고 있었으며 1.0Kb 정도의 EcoR I-HindIII 부분이 더 삽입한 것을 알 수 있었다.

  • PDF

Sex-related demographics in two remnant populations of a dioecious tree Ilex cornuta (Aquifoliaceae): implications for conservation

  • Shin, Sookyung;Lee, Hakbong;Lee, Jei-Wan;Kang, Hyesoon
    • Journal of Ecology and Environment
    • /
    • 제43권3호
    • /
    • pp.320-331
    • /
    • 2019
  • Background: Dioecious plant species having both male and female plants have been investigated regarding sex-related characteristics such as sex ratio, sex-differential resource requirements, and spatial segregation of the sexes. Habitat loss and fragmentation are major threats to the survival of plant populations, but dioecious species are particularly more prone to such habitat degradation than non-dioecious species because of their dimorphic sexual system. We examined the sex-related demographics of two Ilex cornuta populations being different regarding land use history. Methods: During 2016-2017, we examined I. cornuta trees with a basal diameter ${\geq}1.5cm$ in the Yongsu-ri population (YS population) and the Gotjawal Provincial Park population (GP population). Plant sex (male, female, or unsexed) was identified. The tree size (basal diameter and height of the main stem), clonal production (the ramet numbers per genet), and vitality for each clone were measured. The associations between population, sex, tree size, clonal production, and vitality were examined using ANOVAs and contingency table analyses. Finally, point pattern analyses using O-ring statistics were conducted to assess spatial patterns. Results: Upon excluding unsexed trees, the YS population with 74 trees was significantly male-biased (0.66), while the GP population with only 26 trees had a 1:1 sex ratio. In both populations, males and females did not differ in tree size. Although the mean number of ramets differed significantly between populations, females tended to produce more ramets than males. The proportion of weak trees was significantly higher in the YS than in the GP population. Neither population showed evidence of spatial segregation of the sexes. Conclusions: The two populations of dioecious I. cornuta are characterized by the small number of trees and relatively high frequencies of non-reproductive trees. Both indicate that these populations are quite susceptible to environmental and genetic stochasticity. On the other hand, the differences between populations in sex ratio, clonal production, and vitality suggest that conservation efforts for I. cornuta need to be population-specific. In order to help recover and enable this vulnerable species to persist, it is necessary to find ways to enhance their sexual reproduction and simultaneously reduce habitat disturbances due to anthropogenic activities.

Cloning and Characterization of the Lactococcus lactis subsp. lactis ATCC 7962 pts HI Operon

  • Kim, Tea-Youn;Park, Rae-Jun;Chang, Hae-Choon;Chung, Dae-Kyun;Lee, Jong-Hoon;Lee, Hyong-Joo;Kim, Jeong-Hwan
    • Journal of Microbiology and Biotechnology
    • /
    • 제10권6호
    • /
    • pp.829-835
    • /
    • 2000
  • The ptsH and ptsI genes of Lactococus lactis subsp. lactis ATCC 7962 (L. lactis 7962), encoding the general proteins of phosphotransferase system (PTS) components, HPr and enzyme I, respectively, were cloned and characterized. A 1.3 kb PCR product was obtained using a primer set that was hybridized to the internal region of the L. lactis 7962 pts HI genes and then subcloned into a low-copy number vector, pACYC184. The 5' upstream and 3' downstream region from the 1.3 kb fragment were subsequently clone using the chromosome walking method. The complete ptsHI operon was constructed and the nucleotide sequences determined. Two ORFs corresponding to HPr (88 amino acids) and enzyme I (575 amino acids) were located. The ptsHI genes of L. lactis 7962 showed a very high homology (84-90%) with those genes from other Gram-positive bacteria. A primer extension analysis showed that the transcription started at either one of two adjacent bases upstream of the start codon. Using a Northern analysis, two transcripts were detected; the first, a 0.3 kb transcript corresponding to ptsH and the second, a 2 kb transcript corresponding to ptsH and ptsI. The transcription level of ptsH was higher than that of ptsI. The concentration of the ptsH transcript in cells grown on glucose was similar to that in cells grown on lactose, yet higher than that in cells grown on galactose. The ptsI transcript was scarcely detected in cell grown on lactose or galactose. The ptsI transcript was scarcely detected in cells grown on lactose or galactose. The results of a sequence analysis and Northern blot confirmed that the ptsH and ptsI genes of L. lactis 7962 were arranged in an operon like other known ptsHI genes and the expression of the ptsHI genes was regulated at the transcriptional level in response to the carbon source.

  • PDF

Pseudomonas sp. PY002에서 Exotoxin A의 생성에 미치는 철 이온의 영향과 Exotoxin A 유전자의 클로닝 (Effect of Ferrous Ion on the Formation of Exotoxin A from Pseudomonas sp. PY002 and Cloning of it's Gene)

  • 최선아;김호상;최지영;강정숙;김춘성;김덕례;김영주;여명구;박열
    • 미생물학회지
    • /
    • 제35권1호
    • /
    • pp.7-12
    • /
    • 1999
  • Pseudomons sp. PY002의 exotoxin A 의 발현 양상을 관찰하기 위하여 P.aeruginosa PAO1의 anti-exotoxin A와 immunoblot hybridization을 실시한 결과 배지내에 유용 가능한 철이 고갈됨에 따라 exotoxin A의 발현양은 점차적으로 증가하는 양상을 보였으며, CAS 배지에 점적한 배양 상층액에서 siderophore의 발현양도 증가함을 보였다. P.sp.PY002 의 genomix library를 제조하여, exotoxin A를 분비하는 2개의 클론을 선별하려 pETA23과 pETA42 로 명명한 후, 반응성이 강한 Peta42를 선발하였다. pETA 42는 약 1.7kb 크기의 insert를 가지며, 양쪽 말단에 cloning site 인 pstI site 가 존재하며 2개의 NcoI, 1개의 PvuII, 1개의 SstI , 3개의 SmaI, 1개의 KpnI, 3개의 HaeII, 1개의 EcoRI site가 존재하였다.

  • PDF

Cloning and Analysis of a Type II Polyketide Synthase Gene Cluster from Streptomyces toxytricini NRRL 15,443

  • Yoo An-Na;Demirev Atanas V.;Lee, Ji-Seon;Kim, Sang-Dal;Nam Doo-Hyun
    • Journal of Microbiology
    • /
    • 제44권6호
    • /
    • pp.649-654
    • /
    • 2006
  • A standard type II polyketide synthase (PKS) gene cluster was isolated while attempting to clone the biosynthetic gene for lipstatin from Streptomyces toxytricini NRRL 15,443. This result was observed using a Southern blot of a PstI-digested S. toxytricini chromosomal DNA library with a 444 bp amplified probe of a ketosynthase (KS) gene fragment. Four open reading frames [thioesterase (TE), $\beta$-ketoacyl systhase (KAS), chain length factor (CLF), and acyl carrier protein (ACP)], were identified through the nucleotide sequence determination and analysis of a 4.5 kb cloned DNA fragment. In order to confirm the involvement of a cloned gene in lipstatin biosynthesis, a gene disruption experiment for the KS gene was performed. However, the resulting gene disruptant did not show any significant difference in lipstatin production when compared to wild-type S. toxytricini. This result suggests that lipstatin may not be synthesized by a type II PKS.