• Journal of Genetic Medicine
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• 제1권1호
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• pp.45-50
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• 1997

Neuroblastoma, a pediatric malignant neoplasm of neural crest origin, has a wide range of clinical virulence. The mechanisms contributing to the development of neuroblastomas are largely unclear, but non-random chromosomal changes identified over the past years suggest the involvement of genetic alterations. Amplification of the human N-myc proto-oncogene is frequently seen either in extrachromosomal double minutes or in homogeneously staining regions (HSRs) of aggressively growing neuroblastomas. N-myc maps to chromosome 2 band 24, but HSR have never been observed at this band, suggesting transposition of N-myc during amplification. We have constructed and analyzed the region-specific painting probe for HSR in neuroblastoma IMR-32 to determine the derivative chromosomes. Microdissection was performed on HSR using an inverted microscope with the help of microglass needles and an micromanipulator. We pretreated the microdissected fragments with Topoisomerase I which catalyzes the relaxation of supercolled DNA, and performed two initial rounds of DNA synthesis with T7 DNA polymerase followed by conventional PCR to enable the reliable preparation of Fluorescent in situ hybridization probe from a single microdissected chromosome. With this method, it was possible to construct the region-specific painting probe for HSR. The probe hybridized specifically to the HSRs of IMR-32, and to 2p24, 2p13 of normal chromosome. Our results suggest there was coamplification of N-myc together with DNA of the chromosome 2p24 and 2p13. Moreover, the fluorescent signals for the amplified chromosomal regions in IMR-32 cells were also easily recognized at a Thus this painting probe can be applied to detect the similar amplification of N-myc in neuroblastoma tissue, and the probe pool for HSR may be used to identify the cancer-relevant genes.

• 한국연초학회지
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• 제20권1호
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• pp.66-70
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• 1998

TMV resistant lines (TRLs) originated from the Blo plant of Nicotiana tabacum cv. NC82 transformed with TMV coat protein cDNA which initially showed delayed disease symptom were selected for increased resistance in each subsequent generation. The result of field experiment of the transgenic tobacco lines in the fifth generation for TMV resistance and their response to other tobacco diseases (black shank, bacterial wilt, and powdery mildew) is described in this report. When fifteen TRLs of the fifth generation were tested for TMV resistance by mechanically inoculating the individual plants, over 95 percent of the plants of 6 lines showed complete resistance even 8 weeks after the inoculation. Average frequency of the resistant plants in TRLs of the fifth generation 8 weeks after the inoculation was 87%. Stable insertion and expression of TMV coat protein cDNA in the fifth generation of the transgenic tobacco plant were confirmed by PCR and immunoblot hybridization, respectively. All TRLs were resistant to the black shank but were susceptible to the bacterial wilt disease and the powdery mildew to the same degree as non-transgenic NC82 was. Therefore, it was indicated that the phenotypes related at least to disease resistance were not changed in the transgenic tobacco. Key words : TMV CP cDNA, TMV resistant tobacco plant, transformation.

• 한국약용작물학회지
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• 제11권4호
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• pp.311-315
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• 2003

This study was carried out to compare chromosomal characteristics between Atractylodes japonica and A macrocephala. Cytogenetic analysis was conducted based on karyotype analysis and physical mapping using fluorescence in situ hybridization. As a result of karyotype analysis by feulgen staining, somatic chromosome numbers of A. japonica and A. macrocephala were 2n=24. The length. of the mitotic metaphase chromosomes of A. japonica ranged from $0.70\;to\;1.60{\mu}m$ with a total length. of $12.11{\mu}m$ and the homologous chromosome complement comprised six metacentrics, five submetacentrics and one subtelocentrics. On the other hand, the length of the mitotic metaphase chromosomes of A. macrocephala ranged from $0.90\;to\;2.35{\mu}m$ with a total length of $16.58{\mu}m$ and the homologous chromosome complement comprised seven metacentrics and five submetacentrics. The total length of A. japonica chromosomes was shorter than that of A. macrocephala, but A. japonica had one subtelocentrics (chromosomes 4) different from A. macrocepha1a. chromosomes. The F1SH technique using 17S and 5S rDNA was applied to metaphase chromosomes. The signals for 17S rDNA were detected on the telomeric regions of chromosomes 4 and 5 in both A japonica and A. macrocephala. The 5S rDNA signal was found in the short arm of chromosome 1.

• 한국식물학회:학술대회논문집
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• 한국식물학회 1999년도 제13회 식물생명공학심포지움 New Approaches to Understand Gene Function in Plants and Application to Plant Biotechnology
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• pp.17-20
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• 1999

In rice, limited efforts have been made to identify genes by the use of insertional mutagens, especially heterologous transposons such as the maize Ac/Ds. We constructed Ac and gene trap Ds vectors and introduced them into the rice genome by Agrobacterium-mediated transformation. In this report, rice plants that contained single and simple insertions of T-DNA were analyzed in order to evaluate the gene-tagging efficiency. The 3'end of Ds was examined for putative splicing donor sites. As observed in maize, three splice donor sites were identified at the 3'end of the Ds in rice. Nearly 80% of Ds elements wered excised from the original T-DNA sites, when Ac cDNA was expressed under a CaMV 35S promoter. Repetitive ratoon culturing was performed to induce new transpositions of Ds in new plants derived from cuttings. About 30% of the plants carried at least one Ds that underwent secondary transposition in the later cultures. 8% of transposed Ds elements expressed GUS in various tissues of rice panicles. With cloned DNA adjacent to Ds, the genomic complexities of the insertion sites were examined by Southern hybridization. Half of the Ds insertion sites showed simple hybriodization patterns which could be easily utilized to locate the Ds. Our data demonstrate that the Ac/Ds mediated gene trap system could prove an excellent tool for the analysis of functions of genes in rice. We discuss genetic strategies that could be employed in a largee scale mutagenesis using a heterologous Ac/Ds family in rice.

• Journal of Microbiology and Biotechnology
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• 제16권10호
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• pp.1537-1543
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• 2006

A bacterial strain named HY-3 that produces a highly active extracellular protease was isolated from the digestive tract of a spider, Nephila clavata. The bacterium was a Gram-negative, oxidase-negative, catalase-positive, nonhalophilic, nitrate-reducing, facultative anaerobe. Transmission and scanning electron microscopies demonstrated that the isolate was non-spare-forming, straight, rod-shaped, and motile by peritrichous flagella. The G+C content of the DNA was 57.0 mol%. The isoprenoid quinone type was ubiquinone with 8 isoprene units (Q-8). The morphological and biochemical characteristics including the predominant fatty acid and phospholipids profiles placed the isolate HY-3 in the family Enterobacteriaceae. Further biochemical characterization and phylogenetic studies including determination of an almost complete 16S ribosomal DNA sequence suggested that the bacterium was closely related to the genus Serratia. DNA-DNA hybridization analysis revealed that this extracellular protease-producing strain belongs to Serratia proteamaculans, which is also known far its association with insects.

• 한국잠사곤충학회지
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• 제50권2호
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• pp.189-193
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• 2012

To find diapause-related genes, we were performed by differential hybridization with three types of [${\alpha}-^{32}P$]dCTP-labeled total cDNA probes synthesized from diapause-prepared, diapause-maintained and diapause-activated stage of Bombus ignitus queen. Nine individual cDNA clones were found to be differentially expressed in diapause-maintained and diapause-activated stage. Among these clones, BIDC9(BIDC ; Bombus ignitus differentially expressed clone) was analyzed through full-length sequencing and expression pattern analysis. This clone was specifically expressed in the thorax organ. The effect of Juvenile hormone analog(JHA) and $CO_2$ treatment was examined. JHA treatment induced the expression of BIDC9 cDNA clone abruptly after 4 day of treatment. $CO_2$ treatment induced also the clone after 2 day of treatment. BIDC9 cDNA was identified as Bombus ignitus diapause gene contained an open reading frame of 1376 bp encoding 255 amino acids.

• Journal of Microbiology and Biotechnology
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• 제33권8호
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• pp.1084-1090
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• 2023

The strain KIST612, initially identified as E. limosum, was a suspected member of E. callanderi due to differences in phenotype, genotype, and average nucleotide identity (ANI). Here, we found that E. limosum ATCC 8486^T and KIST612 are genetically different in their central metabolic pathways, such as that of carbon metabolism. Although 16S rDNA sequencing of KIST612 revealed high identity with E. limosum ATCC 8486^T (99.2%) and E. callanderi DSM 3662^T (99.8%), phylogenetic analysis of housekeeping genes and genome metrics clearly indicated that KIST612 belongs to E. callanderi. The phylogenies showed that KIST612 is closer to E. callanderi DSM 3662^T than to E. limosum ATCC 8486^T. The ANI between KIST612 and E. callanderi DSM 3662^T was 99.8%, which was above the species cut-off of 96%, Meanwhile, the ANI value with E. limosum ATCC 8486^T was not significant, showing only 94.6%. The digital DNA-DNA hybridization (dDDH) results also supported the ANI values. The dDDH between KIST612 and E. callanderi DSM 3662^T was 98.4%, whereas between KIST612 and E. limosum ATCC 8486^T , it was 57.8%, which is lower than the species cut-off of 70%. Based on these findings, we propose the reclassification of E. limosum KIST612 as E. callanderi KIST612.

• 한국잠사곤충학회지
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• 제52권1호
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• pp.39-44
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• 2014

효율적인 형질전환 누에 시스템 구축을 위해서는 새로운 전이인자의 개발과 함께 선발을 위한 마커 유전자 및 transposase 발현을 효과적으로 조절할 수 있는 다양한 유전자 프로모터 개발이 필수적이다. 이와 관련하여 선행연구를 통해 누에 후부실샘으로부터 고발현하는 RNA binding protein-1 homologue(RBP-1) 유전자를 선발한 바 있다. 본 연구에서는 RBP-1유전자의 누에 발육시기별 및 유충 조직별 발현양상을 Northen blot hybridization 방법으로 분석한 결과, RBP-1 유전자는 유충기로부터 번데기 후기까지의 전기간에 걸쳐 발현하였으며, 두부, 표피, 중장, 지방체 및 견사선 등 실험한 모든 유충 조직에서 고발현 하는 것으로 관찰되었다. 또한, 누에 게놈 유전자은행을 제작한 후 RBP-1 cDNA 유전자를 탐침으로 5'-UTR 영역을 클로닝하고 luciferase assay 방법으로 RBP-1 유전자 프로모터의 활성을 분석하였다. 실험 결과, RBP-1 cDNA를 탐침으로 RBP-1 유전자 ORF와 5'-UTR이 포함된 약 1,660 bp 영역의 게놈 유전자를 클로닝하였다. RBP-1 유전자 프로모터 활성검정을 위해 전사 개시점(+ 30)으로부터 상류의 -740 bp 영역을 PCR로 분리한 후 pGL3 basic vector에 도입하여 luciferase 활성 측정을 위한 전이벡터, pGL-RBP1를 제작하였다. 제작된 pGL-RBP1는 곤충 세포주(Sf9)에 transfection 한 후 luciferase 발현량을 측정한 결과, 기존의 BmA3 유전자 프로모터 대비 10% 가량 높은 발현 효율을 확인할 수 있었다.

• 한국미생물·생명공학회지
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• 제45권2호
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• pp.168-177
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• 2017

Strain A28-5는대한민국 제주도 연안의 해수샘플로부터 고체배지내 xylan과 agar를 분해하는 균주로 분리되었다. Strain A28-5는그람 음성균으로 한 개의 polar flagella로 운동성을 갖는 $Na^+$ 이온 요구성 균주로 분석되었다. 또한 ampilcillin과 thiostreptone 등의 항생제에 내성을 보였다. Genome 내 G+C content는 43.96%이고, Menaquinone-7 (MK-7)을 predominant quinone으로 함유하고 있었다. Strain A28-5의 세포벽을 구성하는 주요 지방산은 $C_{16:1}$ ${\omega}7c/iso-C_{15:0}$ 2-OH (23.32%), $C_{16:0}$ (21.83%), $C_{18:1}$ ${\omega}7c$ (17.98%)였다. strain A28-5의 16S rRNA gene sequence는 Catenovulum agarivorans YM01와 가장 높은 상동성(98.94%)을 보였으며, Neighbor-Joining phylogenetic tree 제작을 통해서 Catenovulum agarivorans YM01와 가장 높은 근연관계를 보이는 것을 증명하였다. Catenovulum agarivorans YM01과의 DNA-DNA hybridization 분석을 통하여 A28-5을 Catenovulum 속의 신종으로 분류하여Catenovulum jejuensis A28-5로 명명하였다. 이 균주의 액체배양으로부터 준비된 두 종류의 조효소를 이용한 xylan 또는 agarose와의 효소반응액을 Thin layer chromatography로 분석하여 각각 tetramer와 hexamer의 xylooligosaccharides와 (neo)agarooligosacchardes가 생산되는 것을 확인하였다.

• 생명과학회지
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• 제14권1호
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• pp.21-25
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• 2004

Crystal violet와 malachite green색소 분해에 관여하는 유전자들을 규명하기 위하여 색소분해능을 가진 Citrobacter sp.의 염색체 DNA속에의 transposon 도입에 의해 생성된 무작위 변이주들이 분리되었다. 이들 변이 주들로부터 두가지 색소분해능을 소실한 14개의 변이주들이 선별되었고, 이들로부터 염색체 DNA를 분리하여 EcoRl으로 절단한 후 Tn5 단편을 probe로하여 Southern hybridization을 행한 결과, 염색체 DNA상의 각각 다른 부위에 Transposon이 Single 삽입된 5개의 변이주 (Cmg2, Cmg6, Cmg8, Cmgll, Cmg12)가 최종적으로 분리되었다. 이들 변이주들의 Transposon 삽입부위 주위의 염기서열과 이로부터 유추되는 아미노산서열을 database상에 등록되어 있는 유전자의 염기서열과 단백질의 아미노산 서열에 대한 상동성을 비교한 결과, Cmg2는 대장균 maltose trnasporter (Mal C)이고, Cmg6은 LysR-type 전사조절 단백질이며, Cmg12는 산화환원효소를 코드하는 유전자인 것으로 알려졌고, 나머지 Cmg8과 Cmg11은 아직까지 기능이 알려져 있지 않은 단백질을 코드하는 유전자인 것으로임이 판명되었다.