• Title/Summary/Keyword: T-DNA transfer

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Evaluation of Disease Resistance of a Leaffolder-resistant (Cry1Ac1) Rice Event and Gene Transfer to Plant Pathogens (혹명나방 저항성벼(Cry1Ac1)의 병해 저항성 및 병원균으로의 유전자 전이)

  • Nam, Hyo-Song;Shim, Hong-Sik;Yu, Sang-Mi;Lee, Se-Won;Kwon, Soon-Jong;Kim, Myung-Kon;Lee, Yong-Hoon
    • Research in Plant Disease
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    • v.15 no.3
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    • pp.202-208
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    • 2009
  • The genetically modified leaffolder-resistant (Cry1Ac1) rice plant was evaluated for the changes of resistance by comparing the occurrence of major diseases with a japonica type Korean rice variety, Nakdong which was the mother plant of the transgenic rice event, in greenhouse and field conditions. There was no difference in the occurrence of sheath blight and Helminthosporium blight between the two varieties in the fields. We couldn't find any difference of resistance for fungal blast and bacterial leaf blight by artificial inoculation in greenhouse. There was also no difference in the susceptibility to sheath blight in artificial inoculation tests confirming the results in the fields. The possibility of gene transfer of Bar and Cry1Ac1 from the genetically modified rice plant to naturally infected pathogens such as Fusarium moniliforme and Pyricularia oryzae in the field conditions was tested by PCR. And the possible transfer of those genes by continuous inoculation of Xanthomonas oryzae pv. oryzae and Rhizoctonia solani was also tested. However, we couldn't find any possibility of transfer of the genes in natural and artificial conditions.

Expression of γ-Tocopherol Methyltransferase Transgene Improves Tocopherol Composition in Lettuce (Latuca sativa L.)

  • Cho, Eun Ae;Lee, Chong Ae;Kim, Young Soo;Baek, So Hyeon;de los Reyes, Benildo G.;Yun, Song Joong
    • Molecules and Cells
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    • v.19 no.1
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    • pp.16-22
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    • 2005
  • A cDNA encoding ${\gamma}-tocopherol$ methyltransferase (${\gamma}-TMT$) from Arabidopsis thaliana was overexpressed in lettuce (Latuca sativa L.) to improve the tocopherol composition. Seven lines of lettuce ($T_0$) containing the ${\gamma}-TMT$ transgene were produced by Agrobacterium-mediated transformation. The inheritance and expression of the transgene were confirmed by DNA and RNA gel blot analyses as well as quantification of tocopherols and ${\gamma}-TMT$ activities. The ratio of ${\alpha}-/{\gamma}-tocopherol$ content (TR) varied from 0.6 to 1.2 in non-transformed plants, while the $T_0$ plants had ratios of 0.8 to 320. The ratio ranged from 0.4 to 544 in 41 $T_1$ progenies of the $T_0$ transgenic line gTM3, and the phenotypic segregation indicated monogenic inheritance of the transgene (i.e., 3:1 = dominant:wild-type classes). There was a tight relationship between the TR phenotype and ${\gamma}-TMT$ activity, and enzyme activities were affected by the copy number and transcript levels of the transgene. The TR phenotype was stably expressed in $T_2$ progenies of $T_1$ plants. The results from this study indicated that a stable inheritance and expression of Arabidopsis ${\gamma}-TMT$ transgene in lettuce results in a higher enzyme activity and the conversion of the ${\gamma}-tocopherol$ pool to ${\alpha}-tocopherol$ in transgenic lettuce.

Genetic Transformation of the Mycelia of Tremella fuciformis and Changes of Cytotoxicity (흰목이 균사체 형질전환 및 세포독성의 변화)

  • Shin, Dong-Il;Park, Hee-Sung
    • The Korean Journal of Mycology
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    • v.41 no.4
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    • pp.287-291
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    • 2013
  • Tremella fuciformis, as one of higher basidiomycetes, can asexually reproduce yeast-like conidium (YLC) cells by budding. We have developed an efficient method to introduce pCambia1300 plasmid containing hph gene into YLC cells using Agrobacterium. This was successful only when YLC cells were wounded by NaOH treatment before co-cultivation. In average, 40~50 transformants were produced out of $1.0{\times}10^6$ YLC cells investigated. The T-DNA transfer was confirmed by PCR. Methanolic extracts from transformants demonstrated different levels of toxicity against SKOV-3 cervical cancer cells.

Molecular cloning of a rhoptry protein (ROP6) secreted from Toxoplasma gondii

  • Ahn Hye-Jin;Kim Seh-Ra;Nam Ho-Woo
    • Parasites, Hosts and Diseases
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    • v.44 no.3
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    • pp.251-254
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    • 2006
  • Monoclonal antibody (mAb) Tg786 against Toxoplasma gondii has been found to detect a 42-kDa rhoptry protein (ROP6) which showed protease activity and host cell binding characteristics after secretion. Using the mAb, a colony containing a 3'-UTR was probed in a T. gondii cDNA expression library. A full length cDNA sequence of the rhoptry protein was completed after 5'-RACE, which consisted of 1,908 bp with a 1,443 bp ORF. The deduced amino acid sequence of ROP6 consisted of a polypeptide of 480 amino acids without significant homology to any other known proteins. This sequence contains an amino terminal stop transfer sequence downstream of a short neutral sequence, hydrophilic middle sequence, and hydrophobic carboxy terminus. It is suggested that the ROP6 is inserted into the rhoptry membrane with both N- and C-termini.

The Complete Mitochondrial Genome of the Fourhorn Sculpin Triglopsis quadricornis (Perciformes, Cottidae) from Sirius Passet, North Greenland

  • Kim, Bo-Mi;Kihm, Ji-Hoon;Park, Tae-Yoon S.
    • Ocean and Polar Research
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    • v.43 no.4
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    • pp.371-374
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    • 2021
  • Triglopsis quadricornis Linnaeus, 1758 (Cottidae) is distributed in the Atlantic and Arctic and has four unique bony protuberances on its head. Here, we report the complete, circular, and annotated mitochondrial genome of T. quadricornis. The complete T. quadricornis mitochondrion was sequenced by high-throughput Illumina HiSeq platform. The sequences are 16,736 bp in size and contains 13 protein-coding genes (PCGs), 22 transfer RNA (tRNA) genes, a control region, and large and small ribosomal subunits. The overall genomic structure of T. quadricornis mitochondrion was conserved with the gene arrangement of Megalocottus and Myoxocephalus species, and phylogenetic analysis supports their sister relationships. Most PCGs consist of TAA or TAG as a termination codon, whereas COII, ND4, and CYTB have T-- as a stop codon. This complete mitochondrial DNA information of T. quadricornis will provide an essential genomic resource to elucidate the phylogenetic relationship and evolutionary history of the family Cottidae.

A Study of the Liposome-Mediated Transgenic Chicken Production (리포좀을 이용한 형질전환 닭 생산에 대한 연구)

  • Byun S. J.;Park C.;Yang B. S.;Kim T. Y.;Sohn S. H.;Kim S. H.;Jeon I. S.
    • Korean Journal of Poultry Science
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    • v.31 no.4
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    • pp.293-298
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    • 2004
  • Microinjection of DNA is a general method for generating transgenic animals, but the rate of transgenesis in chickens is very low. So it was carried out to investigate the efficiency of liposome-mediated gene transfer in stage one cell of chicken embryo with GFP expression vector. In order to determine efficiency and duration of the introduced foreign gene, it was microinjected DNA with liposome or naked DNA into the germinal disc of stage one cell or stage-X chicken embryo. Analysis of reporter gene expression in day-4 embryos showed that GFP expression was observed only in the liposome-mediate embryo groups and detectable up to day-8 embryos. The results suggest that stable integration of the introduced gene using liposome is a rare event. Nevertheless the liposome-mediated gene transfer may be a useful method to transfer a foreign gene into the stage one cell of chicken embryos.

Generation of Transgenic Mice Overexpression Mouse RESISTIN

  • J. R. Chun;S. J. Song;J. T. Do;K. S. Chung;Lee, H. T.
    • Proceedings of the Korean Society of Embryo Transfer Conference
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    • 2002.11a
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    • pp.99-99
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    • 2002
  • The hormone resistin is associated with typeII diabetes mellitus in rodent model. Resistin impairs glucose tolerance and insulin action. A new class of anti-diabetic drugs were called thiazolidinediones (TZDs) downregulates a resistin which is induced during adipocyte differentiation. But the connection between increased adiposity and resistin remains unknown. The objectives of this study was to clone a mouse resistin cDNA and to generate transgenic mice overexpressing mouse resistin gene. The 555 bp of mouse resistin was amplified from mob cDNAS by polymerase chain reaction (PCR) and cloned into pCR$\^$(R)/ 2.1 TOPO T-vector. Mouse resistin mRNA on the basis of Genbank sequence (acession no. AF323080). Then, the PCR product was cloned into pTargeT$\^$TM/ mammalian expression vector that has pCMV promoter and chimeric intron. Restriction enzyme analysis with BamH I and Not I was carried out to determine an orientation of the insert in the vector. The pCMV-mus/resistin gene was prepared from previous recombinant pTargeT$\^$TM/-mus/resistin by digestion of Bgl II, and has used for microinjection into pronuclei of one cell embryos. The microinjected embryos were transfered to pseudopregnant foster-mother. Mouse resistin expression was detected in transgenic F1 mice by Reverse Transcriptase- Polymerase Chain Reaction (RT-PCR). Resistin gene expression mouse has heavier body weight which was measured higher level of plasma glucose than that of normal mouse. And in diet-induced experiments, the abdominal fat pads were isolated from each 24h starvation and re-feeding after fasting group mice that were assessed by RT-PCR analysis. In fasting group mice, resistin expression was higher than that of re-feeding group mice. This result suggests that the resistin gene overexpressing mice may be became to obesity and be useful as an animal disease model to be diabetes mellitus caused by insulin resistance of resistin.

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Specific Localization of DNMT1 in Mouse and Bovine Preimplantation Embryos

  • Y.M.Chang;Min, K.S.;Yoon, J.T.;M.G.Pang;Chung, Y.C.;Kim, C.K.
    • Proceedings of the Korean Society of Developmental Biology Conference
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    • 2003.10a
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    • pp.81-81
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    • 2003
  • DNA methylation is a covalent modification of DNA that can modulate gene expression and is now recognized as a major component of the epigenome. During evolution, the dinucleotide CpG has been progressively eliminated from the genome of higher eukaryotes and is present at only 5% to 10% of its predicted frequency. Approxymately 80% of the remaining CpG sites contain methylated cytosines in most vertebrates and they are distributed in a pattern that is unique in each tissue and is inversely correlated with gene expression. The pattern of methylation is faithfully maintained during cell division by the enzyme Dnmt1, the maintenance DNA methyltransferase, which catalyzes the transfer of a methyl group from S-adenosyl-methionine to the 5'-position of the cytosine ring. We have been identified bovine Dnmt1 cDNA full-length recently (AY173048) Little is known on the functions of Dnmt1 in bovine preimplantation embryos. Thus, we analyzed the specific pattern of Dnmt1 in in vitro derived/nuclear transfer bovine and in vivo derived mouse embryos to monitor the epigenetic reprogramming process. We investigated these process by using indirect immunofluresence with an antibody to Dnmt1. According to other studies, Dnmt1 accumulates in nuclei of early growing oocytes but is sequestered in the cytoplasm of mature oocytes. In 2-cell and 4-cell embryos, Dnmt1 is cytoplasmic, but at the 8-cell stage, it is present only in the nucleus. By the blastocyst stage, Dnmt1o is again found only in the cytoplasm. Thus, nuclear localization of Dnmt1o in preimplantation embryos is limited to the 8-cell stages After implantation, Dnmt1 is localized in the nucleus in mouse. However, we have found different patterns of Dnmt1 nuclear localization. Though we used the common antibody, immune-localization data revealed that Dnmt1 antibody have been detected at the nucleus in 1-cell to blastocyst embryos. Therefore, maybe we think that the functions of Dnmt1 between bovine and mice are different. In order to Identify the mechanisms that regulate DNA methylation in bovine preimplantation embryo, we have plans on using bovine oocyte and somatic specific Dnmt1 antibodies.

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The Complete Chloroplast DNA Sequences of Viola selkirkii (뫼제비꽃(Viola selkirkii)의 엽록체 DNA 염기서열 분석)

  • Ah-Reum Go;Yun-Sun Lee;Kyung-Ah Kim;Kyeong-Sik Cheon;Ki-Oug Yoo
    • Proceedings of the Plant Resources Society of Korea Conference
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    • 2020.12a
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    • pp.55-55
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    • 2020
  • 뫼제비꽃(Viola selkirkii)의 엽록체 DNA 염기서열을 차세대염기서열분석법(NGS)을 이용하여 분석하였다. 재료는 강원도 화천군 일산과 제주도 한라산의 2개체를 사용하였다. 분석결과, 염기서열의 길이는 일산의 뫼제비꽃이 156,774 bp (GC content: 36.30%), 한라산의 뫼제비꽃이 157,451 bp(GC content: 36.30%)로 한라산 개체가 길게 분석되었다. 구간별로 LSC(Large single copy)지역은 한라산 개체(85,950 bp)가 일산 개체(85,930 bp)보다 20 bp 길었으며, SSC(Small single copy)지역은 한라산 개체(17,261 bp)보다 일산 개체가 17,982 bp로 길게 분석되었다. IR(Inverted repeat)지역은 한라산 개체가 27,120 bp로 일산 개체(26,431 bp)보다 길게 분석되었다. 이러한 염기서열 길이의 차이는 종내 개체 간 빈번하게 발생하는 현상으로 IGS와 intron 구간에서 확인 된 단순반복서열의 일부 누락과 IR지역 내의 수축과 확장에 의한 것으로 판단된다. 뫼제비꽃 2개체의 엽록체 게놈을 구성하는 유전자 수는 총 111개로 동일하였으며, protein coding gene 77개, tRNA(transfer RNA) gene 30개, 그리고 rRNA (ribosomal RNA) gene 4개로 구성되어 있었다. 이는 기 발표된 엽록체 DNA 전체 염기서열이 밝혀진 제비꽃속 (Viola) 종류들과 동일한 결과이다.

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Overexpression of Gene Encoding Tonoplast Intrinsic Aquaporin Promotes Urea Transport in Arabidopsis

  • Kim, Sun-Hee;Kim, Kang-Il;Ju, Hyun-Woo;Lee, Ho-Joung;Hong, Suk-Whan
    • Journal of Applied Biological Chemistry
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    • v.51 no.3
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    • pp.102-110
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    • 2008
  • Complementation assay of the urea uptake-defective yeast mutants led to the identification of the Arabidopsis AtTIP4;1 gene encoding the aquaporin. However, its physiological functions still remain elusive. In the present study, histochemical and genetic analyses were performed to understand the physiological roles of AtTIP4;1 in urea uptake. The AtTIP4;1 product was detectible in the roots, but not in the leaves, the stem, and the flower. Its promoter allowed the expression of the $\beta$-glucuronidase reporter gene in the roots and the apical meristem in Arabidopsis. The AtTIP4;1 products were induced under nitrogen-deficient conditions. To investigate the role of the tonoplast intrinsic protein in urea transport and developments, Arabidopsis with the loss- and the gain-of-function mutations by T-DNA insertion in AtTIP4;1 and 35S promoter-mediated overexpression of AtTIP4;1 were identified, respectively. The transfer DNA insertion and the AtTIP4;1-overexpressed plants showed normal growth and development under normal or abiotic stress growth conditions. The urea-uptake studies using $^{14}C$-labeled urea revealed higher accumulation of urea in the AtTIP4;1-overexpressed plants. These results provide evidence that overexpression of AtTIP4;1 leads to the increase in the urea-uptake rate in plants without detectable defects to the growth and development.