• 제목/요약/키워드: DNA separation

검색결과 122건 처리시간 0.035초

닭에서 원시생식세포의 효율적 분리 및 외래 유전자 전이에 관한연구 (Studies on the Efficient Separation of Primordial Germ Cells and Introduction of Foreign DNA in the Chicken)

  • 정동기;한재용
    • 한국가금학회:학술대회논문집
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    • 한국가금학회 1999년도 제16차 정기총회및학술발표회
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    • pp.11-33
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    • 1999
  • This study was conducted to determine the embryonic stages for the isolation of the highest number of PGCs and to improve PGCs enrichment method. The primordial germ cells(PGCs) from different sources of chick embryos were isolated. The embryonic stage having the highest number of PGCs from each sources was selected ; 1-day-old embryos for germinal crescent (stage 6-8), 2.5-day-old embryos for blood (stage 17-18) and 5.5-day-old embryos for gonad (stage 27-28). The number of PGCs from one embryonic germinal crescent, blood and gonad was about 87$\pm$1.8, 103$\pm$4.0, and 932$\pm$10.9, respectively. The viability of PGCs after Ficoll from each sources was similar, showing approximately 70%. the PGCs enrichment method was improved using Ficoll density gradient centrifugation. After this step the purity of PGCs from germinal crescent, blood, and gonad was 45$\pm$9.10%, 85$\pm$1.18%, and 86$\pm$0.19%, respectively. Also, PGCs were picked up by mouth pipette to improve the purity. This improved method for the separation of PGCs from different sources will serve as a useful too to preserve the foundation stocks of poultry and to produce germline chimeras.

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미토콘드리아 DNA CYTB 유전자 서열에 대한 분자 계통과 PCR-RFLP 반수체형에 근거한 제주재래돼지의 모계 기원 (Maternal Origins of the Jeju Native Pig Inferred from PCR-RFLP Haplotypes and Molecular Phylogeny for Mitochondrial DNA CYTB Gene Sequences)

  • 한상현;고문석;정하연;이성수;오홍식;조인철
    • 생명과학회지
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    • 제21권3호
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    • pp.341-348
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    • 2011
  • 제주재래돼지의 모계 혈통에 대한 보다 명확한 이해를 얻기 위해, 본 연구에서는 제주재래돼지의 미토콘드리아 DNA (mtDNA) CYTB 유전자를 분석하고 이를 타 품종들에서 얻은 결과들과 비교하였다. 제주재래돼지를 포함한 돼지 6 품종에서 PCR-RFLP 분석을 수행하였고, RFLP 양상은 돼지 품종들을 뚜렷하게 구분되는 두 가지 반수체형(mtCYTB1 and mtCYTB2)으로 분리시켰다. 제주재래돼지 CYTB 서열들은 계통수 상에서 유럽과 아시아품종 cluster에서 모두 발견되었다. 제주재래돼지 CYTB들 중에서 J2 group은 중국재래돼지품종들과 근연이면서 아시아 고유 돼지 계통들과 함께 출현하였으며, 다른 한 group인 J1에 해당하는 서열들은 유럽돼지 계통들과 함께 위치하였고, 아시아 품종들보다는 스페인의 Iberian 재래돼지들과 근연인 것으로 확인되었다. 이 결과들은 현재 제주도에서 사육되고 있는 제주재래돼지 품종의 모계 기원은 크게 아시아계 돼지와 유럽계 돼지인 것으로 추정됨을 보여준다. 따라서 본 연구결과들은 제주재래돼지 집단은 과거에 가축화된 아시아 고유 돼지품종들과 공통 선조를 공유하고, 또한 20세기에 유입된 유럽계 돼지 품종들도 현재의 집단 형성에 기여한 것임을 시사하고 있다.

Mitotic Cohesin Subunit Mcd1 Regulates the Progression of Meiotic Recombination in Budding Yeast

  • Lee, Min-Su;Yoon, Sang-Wook;Kim, Keun Pil
    • Journal of Microbiology and Biotechnology
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    • 제25권5호
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    • pp.598-605
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    • 2015
  • The cohesin complex holds sister chromatids together and prevents premature chromosome segregation until the onset of anaphase. Mcd1 (also known as Scc1), the α-kleisin subunit of cohesin, is a key regulatory subunit of the mitotic cohesin complex and is required for maintaining sister chromatid cohesion, chromosome organization, and DNA repair. We investigated the function of Mcd1 in meiosis by ectopically expressing Mcd1 during early meiotic prophase I in Saccharomyces cerevisiae. Mcd1 partially regulated the progression of meiotic recombination, sister chromatid separation, and nuclear division. DNA physical analysis during meiotic recombination showed that Mcd1 induced double-strand breaks (DSBs) but negatively regulated homologous recombination during DSB repair; Mcd1 expression delayed post-DSB stages, leading to inefficiencies in the DSB-to-joint molecule (JM) transition and subsequent crossover formation. These findings indicate that meiotic cells undergo Mcd1-mediated DSB formation during prophase I, and that residual Mcd1 could regulate the progression of JM formation during meiotic recombination.

모성 및 사회성 분리 백서 모델의 해마에서 유전자 칩을 이용한 유전자 발현 연구 (Microarray Analysis of Gene Expression in Rat Hippocampus of Maternal Social Separation Model)

  • 이희제;손창희;곽형렬;이상현;한윤희;김수영;박종익;전완주;김성수
    • 생물정신의학
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    • 제13권2호
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    • pp.110-116
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    • 2006
  • Objectives : Alteration of hippocampus was demonstrated in the maternal social separation(MSS) pups, separated from dams on postnatal day(pnd) 14 and placed alone. Therefore, to understand the molecular events involved in the MSS, we have initiated a search for gene profiles that are up or down-regulated in the hippocampus of MSS pups. Methods : Analysis of cDNA microarray was performed by using total RNA extracted from the hippocampus of control and MSS pups on pnd 17. Also, passive-avoidance test was demonstrated on pnd 35. Results : Up-regulation of Nedd4a was observed in the hippocampus of MSS pups. Also, MSS rats showed less elongation of latency in passive avoidance test. Conclusion : We suggest that environmental effects of MSS may be altered the neural and/or glial differentiation and synapse formation-related genes which may lead cognitive alterations in MSS rats.

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Fusarium oxysporum f. sp. lycopersici의 Electrophoretic Karyotype (Electrophoretic Karyotypes of Fusarium oxysporum f. sp. lycopersici)

  • 김영태;김홍기
    • 한국균학회지
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    • 제27권2호통권89호
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    • pp.112-118
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    • 1999
  • 한국, 일본 그리고 미국 등지에서 수집된 Fusarium oxysporum f. sp. lycopersici의 electrophoretic karyotype(EK)을 분석하고자 CHEF-DRII pulsed field gel electrophoresis system(Bio-Rad Laboratories, Melville, NY)으로 각 공시균의 chromosome sized DNA를 분리하였다. EK 분석에 적합한 CHEF gel electrophoresis 조건을 얻기 위해 전기영동 시간 및 전압 그리고 switching interval 등의 조건을 다양하게 바꾸어 가며 실험하였다. 그 결과 국내 균주에서 $0.76{\sim}6.41\;Mb$에 달하는 $9{\sim}11$개의 chromosome sized DNA가 분리되었으며 그 total genome size는 $35.29{\sim}38.92\;Mb$ 이었다. 또한 일본과 미국 균주로 부터 $1.24{\sim}6.85\;Mb$범위의 $9{\sim}11$개의 chromosome sized DNA가 분리되었고 그 total genome size는 $35.32{\sim}43.87\;Mb$ 이었다. 이와 같이 얻어진 각 공시균주의 EK는 chromosome sized DNA의 length range 및 total genome size에서 국내 균주와 외국 균주간의 차이를 잘 반영하였다. 또한 국내 균주의 chromosomal polymorphism은 그 변이가 적어 서로 동일하거나 유사하였으며 외국 균주와 뚜렷이 다른 chromosomal DNA pattern을 나타냈다.

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Expression of Polyhistidine-Containing Fusion Human HepG2 Type Glucose Transport Protein in Spodoptera Cells and Its Purification Using a Metal Affinity Chromatography

  • 이종기
    • 대한의생명과학회지
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    • 제16권3호
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    • pp.201-206
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    • 2010
  • In order to develop procedures for the rapid isolation of recombinant sugar transporter in functional form from away from the endogenous insect cell transporter, gene fusion techniques were exploited. Briefly, BamH1-digested human HepG2 type glucose transport protein cDNA was first cloned into a transfer vector pBlueBacHis, containing a tract of six histidine residues. Recombinant baculoviruses including the human cDNA were then generated by allelic exchange following transfection of insect cells with wild-type BaculoGold virus DNA and the recombinant transfer vector. Plaque assay was then performed to obtain and purify recombinant viruses expressing the human transport protein. All the cell samples that had been infected with viruses from the several blue plaques exhibited a positive reaction in the immnuassay, demonstrating expression of the glucose transport protein. In contrast, no color development in the immunoassay was observed for cells infected with the wild-type virus or no virus. Immunoblot analysis showed that a major immunoreactive band of apparent Mr 43,000~44,000 was evident in the lysate from cells infected with the recombinant baculovirus. Following expression of the recombinant fusion protein with the metal-binding domain and enterokinase cleavage site, the fusion protein was recovered by competition with imidizole using immobilized metal charged resin. The leader peptide was then removed from the fusion protein by cleavage with porcine enterokinase. Final separation of the recombinant protein of the interest was achieved by passage over $Ni^{2+}$-charged resin under binding conditions. The expressed transport protein bound cytochalasin B and demonstrated a functional similarity to its human counterpart.

미소유체시스템을 위한 실용적인 패키징 기술 (Practical Packaging Technology for Microfluidic Systems)

  • 이환용;한송이;한기호
    • 대한기계학회논문집B
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    • 제34권3호
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    • pp.251-258
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    • 2010
  • 본 논문은 다기능 미소유체시스템의 일체형 패키징을 위한 MSI (microfluidic system interface) 기술을 제안하고, 이를 설계, 제작, 시험 평가하였다. MSI 기술을 통해 플러그 방식의 유체 인터커넥터, 유체제어를 위한 미소밸브, 광학 인터페이스를 위한 광학창을 유체시스템에 일체형으로 쉽게 구현할 수 있었다. MSI 기술의 유용성을 보이기 위해 미소 유전자시료전처리시스템에 적용되었으며, 미소 유전자시료전처리시스템은 세포정제, 세포분리, 세포용해, DNA 고체상추출, 중합효소연쇄반응, 그리고 모세관전기영동 기능으로 구성되었다. 나아가 MSI 기술이 적용된 미소 유전자시료전처리시스템의 DNA 고체상추출 및 중합효소연쇄반응의 실험결과로부터 MSI가 미소유체시스템을 위한 실용적 패키징 기술임이 검증되었다.

Phylogenetic Analysis of the Corticiaceae Based on Gene Sequences of Nuclear 18S Ribosomal DNAs

  • Lee, Seung-Shin;Jung, Hack-Sung
    • Journal of Microbiology
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    • 제35권4호
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    • pp.253-258
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    • 1997
  • The nuclear 18S ribosomal RNA genes of seven corticioid species were sequenced. These sequences were analyzed and compared with those of 24 other species of the order Aphyllophorales and phylogenetic trees were constructed using parsimonious methods. Phylogenetic analyses showed that two species among examined members of the Corticiaceae, Resinicium bicolor and Thanatephorus praticola, are located distantly from the remaining six species. The separation of R. bicolor seems to be kphylogenetically significant because it has very unique cystidia. The independent lineage of T. practicola suggests that it is also phylogenetically distinct because it has unusual features like the homobasidium producing secondary spores and the spetal ultrastructure of pore cap. Furthermore, Auriscalpium vulgare, Bondarzewia berkeleyi, and Heterobasidion annosum from different families of the Aphyllophorales proved to be closely related to the species of the Corticiaceae. They all have amyloid spores and grouped with Aleyrodiscus amorphus, which is a member of the Corticiaceae. The amyloidity of spores seems to be an improtant character throughout the order of the Aphyllophorales.

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Microencapsulation Methods for Delivery of Protein Drugs

  • Yoon Yeo;Namjin Baek;Park, Kinam
    • Biotechnology and Bioprocess Engineering:BBE
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    • 제6권4호
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    • pp.212-230
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    • 2001
  • Recent advances in recombinant DNA technology have resulted in development of many new protein drugs. Due to the unique properties of protein druges, they have to be delivered by parenteral injection Although delivery of protein drugs by other routes, such as pulmonary and nasal routes, has shown some promises, to date most protein drugs are administered by par-enteral routs. For long-term delivery of protein drugs by parenteral administration, they have been formulated into biodegradable microspheres. A number of microencapsulation methods have been developed, and the currently used microencapsulation methods are reviewed here, The microen-capsulation methods have been divided based on the method used. They are: solvent evapora-tion/extraction; phase separation (coacervation);spray drying; ionotropic gelation/polyelectrolyte complexation; interfacial polyumerization and supercritical fluid precipitation. Each method is de-scribed fro its applications, advantages, and limitations.

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Effects of Surface Geometry on Polyelectrolyte Adsorption

  • Park, Young-G.;Kim, Key-Seek;Sung, Ki-Chun
    • 한국응용과학기술학회지
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    • 제17권2호
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    • pp.149-156
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    • 2000
  • For the adsorption of polyelectrolyte at the surface of polyacrylamide gel particle, preferential adsorption of the large polyelectrolyte such as DNA is governed by the surface area of an adsorbent. The adsorption equilibrium constant can be varied by surface geometry of porous polymer, and it can be described as a function of ionic strength and surface area. Physical parameters affecting the adsorption were estimated using the theoretical governing equation of polyelectrolyte which electrophoretically moved along the column, and geometrical surface area was estimated by Waldman-Mayer's physical model. The separation of polyelectrolytes was studied using the physical parameters estimated by ionic strength and surface geometry.