• 한국육종학회지
• /
• 제40권2호
• /
• pp.128-135
• /
• 2008

Herbicide resistance is the most common trait being tested and thus herbicide?resistant genetically modified plants are now the most widely cultivated worldwide. Here we developed herbicide?resistant transgenic Agrostis mongolica Roshev. by employing an efficient Agrobacterium?mediated transformation procedure with 25.2% of transformation efficiency. The identification and employment of regenerable and reproducible type of callus was one of the most critical factors to ensure success in this study. PCR analysis confirmed that the bar transgene was integrated into the genome of transgenic plants. The expression of 35S?bar gene was confirmed by Northern blot analysis. The transgenic plants showed complete resistance to herbicide, indicating that the bar gene is functional in transgenic plants.

• Journal of Plant Biotechnology
• /
• 제2권1호
• /
• pp.35-41
• /
• 2000

Transgenic cabbage (Brassica oleracea ssp. capitata) plants resistant to the commercial herbicide Bast $a^{R}$ were obtained by Agrobacterium tumefaciens - mediated transformation. Hypocotyl segments of in vitro grown plants were infected with Agrobacterium tumefaciens LBA 4404 harboring plasmid pMOG6-Bar which contains hpt and bar genes. Explants were cultured on callus induction medium (MS basal medium + 1 mg/L NAA + 2 mg/L BA + 2 mg/L AgN $O_3$+ 100 mg/L carbenicillin + 250 mg/L cefotaxime) supplemented with 15 mg/L hygromycin. Hygromycin resistant calluses were transferred to shoot regeneration medium (MS basal medium + 0.1 mg/L NAA + 2 mg/L BA + 3% sucrose + 2 mg/L AgN $O_3$+ 15 mg/L hygromycin + 250 mg/L cefotaxime + 100 mg/L carbenicillin). In order to induce roots, elongated shoots were placed on the MS medium without plant growth regulators and hygromycin. Southern blot analysis of several putative transgenic plants indicated that one to five intact copies of Apt and bar genes were incorporated into the genome. Expression of bar gene was confirmed by Northern blot analysis and by herbicide resistant phenotype. Seed progeny from self-pollinated transformants expressed the herbicide resistance and showed Mendelian segregation of the introduced gene.e.

• 생명과학회지
• /
• 제7권3호
• /
• pp.172-175
• /
• 1997

Transgenic hybrid poplar subclones containing herbicide glyphosate resistant gene (aroA) were treated with ozone at the concentration of 100 nL L$^{-1}$ for 6 hr for 5 consecutive days. The foreign gene expression in leaves of all treated plants was reduced both at transcriptional and translational levels confirmed by Northern and Western blot analysis, respectively, as compared to non-treated control plants. These results indicated that the expression of foreign gene in transgenic plants could be affected by the environmental stresses. Thus, the performance of transgenic plants cultivated on field conditions may be lower than they are expected.

• Journal of Plant Biotechnology
• /
• 제31권4호
• /
• pp.261-266
• /
• 2004

Agrobacterium을 이용하여 도입된 유전자의 세대진전, 교잡 등에 따른 유전적 안정성을 확인코자, 형질 전환으로부터 얻어진 bar 유전자가 도입된 형질전환 식물체들을 상호 교배, 여교배, T$_4$ 세대까지의 자식의 반복 등에 의해 유전적 안정성을 검토하였다. 조합이나 계통에 따라서는 일부 멘델식 분리를 따르지 않고 제초제 Basta에 대한 저항성이 사라지거나 저항성개체보다 감수성개체가 기대치보다 많은 등의 경우가 있었으나, 대부분 멘델식 분리를 따르고 있어 세대진전, 교배 등에 의해서도 유전적 안정성이 높게 유지됨을 확인할 수 있었다.

• 한국환경농학회지
• /
• 제31권1호
• /
• pp.52-59
• /
• 2012

BACKGROUND: Cultivation of genetically modified(GM) crops rapidly has increased in the global agricultural area. Among those, herbicide resistant GM crops are reported to have occupied 89.3 million hectares in 2010. However, cultivation of GM crops in the field evoked the concern of the possibility of gene transfer from transgenic plant into soil microorganisms. In our present study, we have assessed the effects of herbicide-resistant GM Chinese cabbage on the surrounding soil microbial community. METHODS AND RESULTS: The effects of a herbicide-resistant genetically modified (GM) Chinese cabbage on the soil microbial community in its field of growth were assessed using a conventional culture technique and also culture-independent molecular methods. Three replicate field plots were planted with a single GM and four non-GM Chinese cabbages (these included a non-GM counterpart). The soils around these plants were compared using colony counting, denaturing gradient gel electrophoresis and a species diversity index assessment during the growing periods. The bacterial, fungal and actinomycetes population densities of the GM Chinese cabbage soils were found to be within the range of those of the non-GM Chinese cabbage soils. The DGGE banding patterns of the GM and non-GM soils were also similar, suggesting that the bacterial community structures were stable within a given month and were unaffected by the presence of a GM plant. The similarities of the bacterial species diversity indices were consistent with this finding. CONCLUSION: These results indicate that soil microbial communities are unaffected by the cultivation of herbicide-resistant GM Chinese cabbage within the experimental time frame.

• 한국자원식물학회지
• /
• 제11권2호
• /
• pp.188-194
• /
• 1998

This experiment was conducted to introduce phosphinothricin acetyl -transferase(PAT) gene, resistant to basta and non-selective herbidide, into tobacco(Nicotiana tabacum cv.BY4). For shoot formation,tobacco leaf disks were placed on the MS medium supplemented with 2.0mg/L BA and 0.1mg/L NAA. In this medium condition, tobacco leaf disces were cocultivated with A. tumefaciens MP90 containing NPT IIand PAT resistant to kanamycin and Basta, respectively. Shoots were obtained in the medium containing antibiotics, and those were transferred to rooting medium supplemented with 0.1mg/L NAA and antibiotics. The plants obtaining roots were transplanted into soil. Phenotype of transgenic tobacco plant was mostly as normal plant. However, about 5% was abnormal plant, which did not set seeds. PCR analysis and southern blot were performed to determine transformation. As the results, it was confirmed that PAT gene was stably integrated into tobacco genome.When herbicide, basta, was sprayed to the plants confirmed by PCR, the transgenic plants showed normal growth, whereas normal plants died. Therefore, the result of this experiment show that tobacco transformation for the resistance to basta, non-selective herbicide, was successful because PAT gene was stably integrated into tobacco.

• Applied Biological Chemistry
• /
• 제37권4호
• /
• pp.248-254
• /
• 1994

Bromoxynil은 쌍떡잎 특이적 제초제로써 폭 넓게 이용되고 있으며 반감기가 매우 짧다. Bromoxynil을 3,5-dibromo-4-hydroxybenzoic acid로 분해하는 nitrilase를 암호화한 bxn 유전자를 식물 벡터인 pGA482에 도입하고 Agrobacterium과의 동시배양을 통해 담배와 상추에 형질전환하였다. Kanamycin을 이용해 형질전환 식물체를 선별하고 완전한 식물체로 분화시켰다. Northern hybridization을 통해 bxn 유전자의 발현정도를 검정하고 liaf-disc와 pot assay를 통해 형질전환 식물체가 고농도의 bromoxynil에 저항성을 보임을 확인하였다.

• BMB Reports
• /
• 제31권3호
• /
• pp.287-295
• /
• 1998

Acetolactate synthase (ALS) is the first common enzyme in the biosynthesis of L-Ieucine, L-isoleucine, and L-valine. The sulfonylurea-resistant ALS gene from Nicotiana tabacum was cloned into the bacterial expression vector pGEX-2T. The resulting recombinant plasmid pGEX-ALS3 was used to transform Escherichia coli strain XL1-Blue, and the mutant tobacco ALS (mALS) was expressed in the bacteria as a protein fused with glutathione S-transferase (GST). The fusion product GST-mALS was purified in a single step on a glutathione-Sepharose column. ALS activities of 0.9-2.5 ${\mu}mol/min/mg$ protein were observed in the GST-mALS, and the Km values for pyruvate, FAD, and TPP were 10.8-24.1, $(1.9-8.9){\times}10^{-3}$, and 0.14-0.38 mM, respectively. The purified GST-mALS was resistant to both the sulfonylurea and the triazolopyrimidine herbicides, and lost its sensitivity to end products, L-valine and L-leucine. For comparision, the tobacco wild-type recombinant ALS fused with GST, GST-wALS, was also characterized with respect to its pyruvate and cofactor bindings. These results suggest that the purified mutant recombinant tobacco ALS was functionally active, that the mutations resulting in herbicide resistance has affected pyruvate and cofactor bindings," and that the two classes of herbicides interact at a common site on the plant ALS.

• Plant Biotechnology Reports
• /
• 제3권4호
• /
• pp.277-283
• /
• 2009

One of the limitations to conducting maize Agrobacterium-mediated transformation using explants of immature zygotic embryos routinely is the availability of the explants. To produce immature embryos routinely and continuously requires a well-equipped greenhouse and laborious artificial pollination. To overcome this limitation, an Agrobacterium-mediated transformation system using explants of type II embryogenic calli was developed. Once the type II embryogenic calli are produced, they can be subcultured and/or proliferated conveniently. The objectives of this study were to demonstrate a stable Agrobacterium-mediated transformation of maize using explants of type II embryonic calli and to evaluate the efficiency of the protocol in order to develop herbicide-resistant maize. The type II embryogenic calli were inoculated with Agrobacterium tumefaciens strain C58C1 carrying binary vector pTF102, and then were subsequently cultured on the following media: co-cultivation medium for 1 day, delay medium for 7 days, selection medium for $4{\times}14$ days, regeneration medium, and finally on germination medium. The T-DNA of the vector carried two cassettes (Ubi promoter-EPSPs ORF-nos and 35S promoter-bar ORF-nos). The EPSPs conferred resistance to glyphosate and bar conferred resistance to phosphinothricin. The confirmation of stable transformation and the efficiency of transformation was based on the resistance to phosphinothricin indicated by the growth of putative transgenic calli on selection medium amended with $4mg\;1^{-1}$ phosphinothricin, northern blot analysis of bar gene, and leaf painting assay for detection of bar gene-based herbicide resistance. Northern blot analysis and leaf painting assay confirmed the expression of bar transgenes in the $R_1$ generation. The average transformation efficiency was 0.60%. Based on northern blot analysis and leaf painting assay, line 31 was selected as an elite line of maize resistant to herbicide.

• 식물조직배양학회지
• /
• 제28권1호
• /
• pp.53-58
• /
• 2001

제초제 저항성 오이 (Cucumis sativus L. cv Green angel)를 생산하기 위하여 배발생 현탁배양세포와 binary vector pGA-bar을 지닌 Agrobacterium tumefacians (LBA4404)를 공동배양하였다. 형질전환 벡터의 T-DNA부분에는 kanamycin에 저항성을 나타내는 neomycin phosphotrans ferase (npt II) 유전자와 phosphinothricin (PPT)에 저항성을 나타내는 phosphinothricin acetyltransferase (bar) 유전자를 지니고 있다. 48시간의 공동배양 후 배발생 캘러스는 20mg/L PPT가 함유된 성숙배지에서 배양하였다. 약 200개체의 형질전환 유식물체를 40mg/L PPT가 첨가된 호르몬이 없는 배지에서 생산하였다. 5개의 오이 형질전환 식물체의 염색체에 bar유전자가 도입되어 발현되는 것을 northern blot 분석을 통하여 확인하였다. 형질전환 오이 식물체가 토양에서 성숙되었다. 성숙한 오이 식물체는 PPT가 함유된 상업적 제초제 (Basta)를 일반적인 사용 농도 (3ml/L)처리시에도 저항성을 나타내며 생장하였다.