• 제목/요약/키워드: tandem structure

검색결과 113건 처리시간 0.021초

효모의 재조합 변이주를 이용한 인간 Centromeric Alphoid DNA Repeat의 안정성에 관한 연구 (Stability of Human Centromeric Alphoid DNA Repeat during Propagation in Recombination-Deficient Yeast Strains)

  • 김광섭;신영선;이상엽;안은경;도은주;박인호;임선희;선우양일
    • 미생물학회지
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    • 제43권4호
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    • pp.243-249
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    • 2007
  • Centromere는 채세포분열과 생식세포분열 등 맡은 주요 기능을 담당하는 고도로 분화된 구조이다. Alphoid DNA (${\alpha}$-satellite)는 인간뿐 아니라 모든 영장류의 염색체 내 centromere에서 발견되는 반복서열의 대부분을 차지한다. 인간 인공염색체(Human Artificial Chromosome, HAC)의 개발에서 가장 핵심적인 부분은 centromere의 분리 및 안정적인 유지에 있다. 이 영역은 출아효모에서 alphoid DNA 반복서열을 hook으로 이용하여 Transformation-associated recombination (TAR) cloning법을 사용하여 선택적으로 분리할 수 있다. 이러한 실험방법으로 먼저 repeat array를 rolling-circle amplication (RCA)를 통하여 약 5 kb까지 길이를 연장시킨 후, 효모내에서 상동성재 조합을 이용한 TAR cloning법을 사용하여 분리할 수 있다. 이렇게 분리된 35 kb-50 kb 길이의 4종류의 centromeric DNA repeat arrays (2,4,5,6 mer)를 사용하여, 반복서열의 안정성 유지를 조사하기 위해 상동성재조 합 변이주인 rad51, rad52, rad54를 사용하여 비교 분석하였다. 야생주, rad51과 rad54 변이주를 이용하여 형질전환을 수행한 결과, 반복서열의 크기에 있어서 많은 변화를 나타내었다. 반면, rad52 변이주는 야생주와 다르게 형질전환빈도가 매우 낮은 비율로 나타났으나, centromeric DNA repeat array의 안정성은 3배 이상으로 높게 나타냈다. 이러한 결과들을 미루어, rad52 변이주를 사용하여 centromeric DNA repeat arrays의 형질전환실험에서 발생하는 맡은 변이를 줄일 수 있을 것으로 보인다. 이러한 유전적 방법은 HAC 제작에서 반복서열의 유지에 훨씬 효율적으로 사용할 수 있을 것으로 사료된다.

mtDNA D-loop의 염기서열에 의한 제주견과 우리나라 재래견 및 외국견품종과의 유연관계 (Phylogenetic Relationships of Jeju Dogs to Other Domestic and Foreign Dog Breeds Determined by Using mtDNA D-loop Sequences)

  • 김미경;김남영;이성수;김규일;양영훈
    • Journal of Animal Science and Technology
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    • 제53권4호
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    • pp.303-310
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    • 2011
  • mtDNA D-loop 970 bp의 염기서열을 이용하여 제주견과 우리나라의 토종견인 진도견, 풍산견, 삽살견과의 관계와 외국 품종들과의 유연관계를 분석하였다. 또한 초가변영역인 598 bp의 D-loop 부위가 보고된 염기서열들은 추가로 GenBank에서 수집하여 전체 30개의 품종으로부터 214종의 단상형들을 이용하여 AMOVA 분석을 하였다. 염기서열 분석결과 제주견에서 5종류, 진도견 4종류, 삽사리 4종류, 풍산견 5종류, 이스트라이카와 웨스트라이카(Canis familiaris) 각 2종류, 회색늑대(Canis lupus) 2종류, 코요테(Canis latrans) 2종류의 단상형을 얻었다. 한국, 일본, 중국, 유럽의 견품종 사이에 지역의 차이로부터 오는 변이성(1.4%)은 동일 지역에 서로 다른 품종들 사이의 변이(16.2%) 보다도 매우 적은 변이를 보이고 있었다. 개의 조상으로 여기는 늑대와 오늘날 개의 집단과의 염기서열 분산성분 분석에서 개(1.63%)와 늑대(3.64%)의 집단내 변이보다는 개와 늑대 사이의 진단간 변이(4.51%)가 높게 나타난 것으로 나타났으며, 이 값을 근거로 하면 개와 늑대사이에 유전적 분기시기는 약 1~2백 만년 전인 것으로 추정되었다. 염기서열의 변이성과 유연관계분석에서 제주견, 진도견, 풍산견, 삽살견 모두 독특한 계통분기를 형성하지 못했다. 즉 이러한 결과는 국내 토종견들 사이와 또는 유입된 외래품종들 사이에 오랜 세월 동안 서로 상당한 교류가 있었을 것으로 생각되었다. 제주견에서는 삽살이, 진도견, 풍산견의 염기서열과 가까운 유연관계를 보이는 단상형들이 산포되어 있는 것으로 확인되어 제주견 집단이 우리나라 재래종들과의 모계조상에 있어서 구분은 어려운 것으로 확인되었다. 제주견이 집단으로 고정정도를 가늠하는 Fixation index인 Fst값은 진도견, 풍산견, 삽살견 가운데 가장 낮게 나타났다.

Brief Introduction of Research Progresses in Control and Biocontrol of Clubroot Disease in China

  • He, Yueqiu;Wu, Yixin;He, Pengfei;Li, Xinyu
    • 한국균학회소식:학술대회논문집
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    • 한국균학회 2015년도 춘계학술대회 및 임시총회
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    • pp.45-46
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    • 2015
  • Clubroot disease of crucifers has occurred since 1957. It has spread to the whole China, especially in the southwest and nourtheast where it causes 30-80% loss in some fields. The disease has being expanded in the recent years as seeds are imported and the floating seedling system practices. For its effective control, the Ministry of Agriculture of China set up a program in 2010 and a research team led by Dr. Yueqiu HE, Yunnan Agricultural University. The team includes 20 main reseachers of 11 universities and 5 institutions. After 5 years, the team has made a lot of progresses in disease occurrence regulation, resources collection, resistance identification and breeding, biological agent exploration, formulation, chemicals evaluation, and control strategy. About 1200 collections of local and commercial crucifers were identified in the field and by artificiall inoculation in the laboratories, 10 resistant cultivars were breeded including 7 Chinese cabbages and 3 cabbages. More than 800 antagostic strains were isolated including bacteria, stretomyces and fungi. Around 100 chemicals were evaluated in the field and greenhouse based on its control effect, among them, 6 showed high control effect, especially fluazinam and cyazofamid could control about 80% the disease. However, fluzinam has negative effect on soil microbes. Clubroot disease could not be controlled by bioagents and chemicals once when the pathogen Plasmodiophora brassicae infected its hosts and set up the parasitic relationship. We found the earlier the pathogent infected its host, the severer the disease was. Therefore, early control was the most effective. For Chinese cabbage, all controlling measures should be taken in the early 30 days because the new infection could not cause severe symptom after 30 days of seeding. For example, a biocontrol agent, Bacillus subtilis Strain XF-1 could control the disease 70%-85% averagely when it mixed with seedling substrate and was drenching 3 times after transplanting, i.e. immediately, 7 days, 14 days. XF-1 has been deeply researched in control mechanisms, its genome, and development and application of biocontrol formulate. It could produce antagonistic protein, enzyme, antibiotics and IAA, which promoted rhizogenesis and growth. Its The genome was sequenced by Illumina/Solexa Genome Analyzer to assembled into 20 scaffolds then the gaps between scaffolds were filled by long fragment PCR amplification to obtain complet genmone with 4,061,186 bp in size. The whole genome was found to have 43.8% GC, 108 tandem repeats with an average of 2.65 copies and 84 transposons. The CDSs were predicted as 3,853 in which 112 CDSs were predicted to secondary metabolite biosynthesis, transport and catabolism. Among those, five NRPS/PKS giant gene clusters being responsible for the biosynthesis of polyketide (pksABCDEFHJLMNRS in size 72.9 kb), surfactin(srfABCD, 26.148 kb, bacilysin(bacABCDE 5.903 kb), bacillibactin(dhbABCEF, 11.774 kb) and fengycin(ppsABCDE, 37.799 kb) have high homolgous to fuction confirmed biosynthesis gene in other strain. Moreover, there are many of key regulatory genes for secondary metabolites from XF-1, such as comABPQKX Z, degQ, sfp, yczE, degU, ycxABCD and ywfG. were also predicted. Therefore, XF-1 has potential of biosynthesis for secondary metabolites surfactin, fengycin, bacillibactin, bacilysin and Bacillaene. Thirty two compounds were detected from cell extracts of XF-1 by MALDI-TOF-MS, including one Macrolactin (m/z 441.06), two fusaricidin (m/z 850.493 and 968.515), one circulocin (m/z 852.509), nine surfactin (m/z 1044.656~1102.652), five iturin (m/z 1096.631~1150.57) and forty fengycin (m/z 1449.79~1543.805). The top three compositions types (contening 56.67% of total extract) are surfactin, iturin and fengycin, in which the most abundant is the surfactin type composition 30.37% of total extract and in second place is the fengycin with 23.28% content with rich diversity of chemical structure, and the smallest one is the iturin with 3.02% content. Moreover, the same main compositions were detected in Bacillus sp.355 which is also a good effects biocontol bacterial for controlling the clubroot of crucifer. Wherefore those compounds surfactin, iturin and fengycin maybe the main active compositions of XF-1 against P. brassicae. Twenty one fengycin type compounds were evaluate by LC-ESI-MS/MS with antifungal activities, including fengycin A $C_{16{\sim}C19}$, fengycin B $C_{14{\sim}C17}$, fengycin C $C_{15{\sim}C18}$, fengycin D $C_{15{\sim}C18}$ and fengycin S $C_{15{\sim}C18}$. Furthermore, one novel compound was identified as Dehydroxyfengycin $C_{17}$ according its MS, 1D and 2D NMR spectral data, which molecular weight is 1488.8480 Da and formula $C_{75}H_{116}N_{12}O_{19}$. The fengycin type compounds (FTCPs $250{\mu}g/mL$) were used to treat the resting spores of P. brassicae ($10^7/mL$) by detecting leakage of the cytoplasm components and cell destruction. After 12 h treatment, the absorbencies at 260 nm (A260) and at 280 nm (A280) increased gradually to approaching the maximum of absorbance, accompanying the collapse of P. brassicae resting spores, and nearly no complete cells were observed at 24 h treatment. The results suggested that the cells could be lyzed by the FTCPs of XF-1, and the diversity of FTCPs was mainly attributed to a mechanism of clubroot disease biocontrol. In the five selected medium MOLP, PSA, LB, Landy and LD, the most suitable for growth of strain medium is MOLP, and the least for strains longevity is the Landy sucrose medium. However, the lipopeptide highest yield is in Landy sucrose medium. The lipopeptides in five medium were analyzed with HPLC, and the results showed that lipopeptides component were same, while their contents from B. subtilis XF-1 fermented in five medium were different. We found that it is the lipopeptides content but ingredients of XF-1 could be impacted by medium and lacking of nutrition seems promoting lipopeptides secretion from XF-1. The volatile components with inhibition fungal Cylindrocarpon spp. activity which were collect in sealed vesel were detected with metheds of HS-SPME-GC-MS in eight biocontrol Bacillus species and four positive mutant strains of XF-1 mutagenized with chemical mutagens, respectively. They have same main volatile components including pyrazine, aldehydes, oxazolidinone and sulfide which are composed of 91.62% in XF-1, in which, the most abundant is the pyrazine type composition with 47.03%, and in second place is the aldehydes with 23.84%, and the third place is oxazolidinone with 15.68%, and the smallest ones is the sulfide with 5.07%.

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