• Journal of Microbiology and Biotechnology
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• v.17 no.4
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• pp.681-684
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• 2007

An immunoassay method was developed to quantitatively detect phosphinothricin-N-acetyltransferase (PAT) encoded by the Bialaphos resistance (bar) gene in genetically modified (GM) pepper. The histidine-tagged PAT was overexpressed in Escherichia coli M15 (pQE3l-bar) and efficiently purified by $Ni^{2+}$ affinity chromatography. A developed sandwich enzyme-linked immunosorbent assay (S-ELISA) method (detection limit: $0.01{\mu}g/ml$) was 100-fold more sensitive than a competitive indirect ELISA (CI-ELISA) method or Western blot analysis in detecting the recombinant PAT. In real sample tests, PAT in genetically modified herbicide-tolerant (GMHT) peppers was successfully quantified [$4.9{\pm}0.4{\mu}g/g$ of sample (n=6)] by the S-ELISA method. The S-ELISA method developed here could be applied to other GMHT crops and vegetables producing PAT.

• Journal of Microbiology and Biotechnology
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• v.17 no.2
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• pp.335-341
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• 2007

For the development of qualitative and quantitative PCR methods of genetically modified (GM) pepper developed in Korea, a capsanthin-capsorubin synthase (CCS) gene was used as the endogenous reference gene. The primer pair ccs-F/R amplifying the pepper endogenous gene gave rise to an amplicon of 102 bp. No amplified product was observed when DNA samples from 16 different plants were used as templates. The construct-specific primer pairs amplifying the junction region of the bar gene and Ti7 introduced in GM pepper gave rise to an amplicon of 182 bp. Quantitative PCR assay was performed using a TaqMan probe and a standard plasmid as a reference molecule, which contained both an endogenous and event-specific sequence. For the validation of this method, the test samples containing 0.1, 1, 3, 5, and 10% GM pepper were quantified.

• The Korean Journal of Community Living Science
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• v.15 no.4
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• pp.177-184
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• 2004

This study performed compositional analyses of a genetically modified herbicide-tolerant red pepper (GMHT), developed by the Rural Development Administration, and a parental red pepper cultivar 'Subicho', and compared the nutrient composition of them. Using the Association of Official Analytical Chemists (AOAC) methods, the study measured the concentration of nutrients, including the proximate components (protein, fat, fiber, ash, and carbohydrates) and minerals (Ca, K, Na, Fe, Mg, Zn) of GMHT and 'Subicho'. Nutritional composition of GMHT and 'Subicho' were compared with the nutritional composition of conventional red peppers. The nutrient composition of GMHT and 'Subicho' were found similar, and GMHT's nutrient contents were in the range of those of the conventional red peppers. These results showed that GMHT's nutrient contents were equivalent to those of the parental red pepper and other conventional red peppers.

• Food Science and Preservation
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• v.22 no.6
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• pp.901-907
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• 2015

Genetically modified (GM) pepper H15 containing the gene for cucumber mosaic virus (CMV) coat protein (CP) and its control line non-GM pepper P2377 were investigated for their allergic risk. Amino acid sequence of the inserted gene product CMV-CP was compared with those of known allergens. No known allergen had greater than 35% amino acid sequence homology over an 80 amino acid window or more than 8 consecutive identical amino acids. Protein patterns of GM and non-GM pepper extracts were evaluated by SDS-PAGE, which showed similar distribution of protein bands for both GM and non-GM pepper. Antigen-antibody reactions were compared between GM and its non-transgenic parental control. ELISA and immunoblot analysis of sera from allergic patients showed some IgE reactivity; however, no differences were observed between GM pepper H15 and P2377. We therefore conclude that CMV-CP is less likely to be an allergen; the protein composition and allergenicity of the GM pepper H15 is not different from that of P2377 and safe as a commercial host.

• Horticultural Science & Technology
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• v.29 no.2
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• pp.123-129
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• 2011

To identify unintended vertical gene-transfer rates from the developed transgenic plants, rapid and unequivocal techniques are needed to identify event-specific markers based on flanking sequences around the transgene and to distinguish zygosity such as homo- and hetero-zygosity. To facilitate evaluation of zygosity, a polymerase chain reaction technique was used to analyze a transgenic pepper line B20 (homozygote), P915 wild type (null zygote), and their F1 hybrids, which were used as transgene contaminated plants. First, we sequenced the 3'-flanking region of the T-DNA (1,277 bp) in the transgenic pepper event B20. Based on sequence information for the 3'- and 5'-flanking region of T-DNA provided in a previous study, a primer pair was designed to amplify full length T-DNA in B20. We successfully amplified the full length T-DNA containing 986 bp from the flanking regions of B20. In addition, a 1,040 bp PCR product, which was where the T-DNA was inserted, was amplified from P915. Finally, both full length T-DNA and the 1,040 bp fragment were simultaneously amplified in the F1 hybrids; P915 ${\times}$ B20, Pungchon ${\times}$ B20, Gumtap ${\times}$ B20. In the present study, we were able to identify zygosity among homozygous transgenic event B20, its wild type P915, and hemizygous F1 hybrids. Therefore, this novel zygosity identification technique, which is based on PCR, can be effectively used to examine gene flow for transgenic pepper event B20.

• Horticultural Science & Technology
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• v.30 no.2
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• pp.192-200
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• 2012

For the development of genetically modified plants, it is important to verify various factors which potentially affect the risk assessment as well as to establish an experimental program to produce scientific and reliable data. However, it is a time and cost consuming process to develop GM plants as well as to prepare scientific and convincible data for government's approval. Therefore, using the transgenic hot pepper tolerant to a new CMV pathotype, we attempted to suggest few methodological procedures, such as probe saturation for southern blot analysis and RT-PCR and ELISA for expression analysis, for identification and stability evaluation of inserted gene in genetically modified plant which are required for submission for approval. Ten partially overlapped probes covering full length of inserted gene were produced. We could identify that the inserted gene was stacked as a single copy as well as no partial element existed. Also, we could identify the stability of the inserted gene stacked in hot pepper using probe saturation. In the expression analysis with RT-PCR and ELISA, we also could provide the stable expression of transcript and proteins in leaves and placenta and pericarp of fruits of the CMV-resistant hot pepper.

• Journal of Environmental Science International
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• v.20 no.9
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• pp.1183-1190
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• 2011

This study was aimed to establish the evaluating protocol and standard assessment for genetically modified (GM) hot pepper and to find out a proper statistic method to analyze for equality of agricultural characters between GM and non-GM pepper lines. GM and non-GM hot pepper lines were cultivated in two GMO fields in the middle region of Korea and total of 52 agricultural characters were collected during the plant growing season for 4 years, 2007 to 2010. Levene's test was conducted to confirm the homogeneity of raw data before statistic analysis. Two-way ANOVA in the multivariate tests and t-test were conducted to analyze 52 agricultural characters in order to find out the equality between H15 and P2377. From the statistical analysis through two-way ANOVA, 16 out of 16 plant growth traits, 9 out of 18 green fruit traits and 7 out of 18 red fruit traits among 4 years and 9 out of 16 plant growth traits, 4 out of 18 green fruit traits and 3 out of 18 red fruit traits between H15 and P2377 have shown the statistic differences. With the same raw data of 52 agricultural characters, t-test was also conducted. Based on the result from t-test, only 1 out of 16 plant growth traits, 2 out of 18 green fruit traits and 1 out of 18 red fruit traits have shown the differences between H15 and P2377, so that it was concluded that there is no statistic difference between H15 and P2377 in terms of agricultural characters. Also, the t-test is a proper statistic method to analyze each trait between GM and its control lines in order to evaluate agricultural characters.

• 식품문화 한맛한얼
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• v.2 no.1
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• pp.5-31
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• 2009

Prior to 1970, it was known that Korean had our own red pepper named as Kochu and we used Kochu in preparing kimchi and kochujang. However, after Professor Lee insisted that Korean red pepper (Kochu) was transferred from Japan during the Seven Years War (Imjinwaeran(壬辰倭亂)), Japanese Invasion of Korea in 1592${\sim}$1598), it has been generally accepted without any criticisms. But many old literatures have shown that Korean Kochu already existed in Korea before the war. For example, the books, Kukubkanibang ((救急簡易方) published in 1489) and Hunmongjahoi ((訓蒙字會) published in 1527), demonstrated that Kochu was cultivated as food substances or medicinal purposes. In another old literature (1460), Siklyochanyo(食療撰要), kochujang was used as an uncomfort-stomach stabilizer. In addition, Korean red pepper was genetically different from South-Mid America's red pepper called as Aji. It has been also insisted by Professor Lee that Aji was transferred to Europe by Columbus in 1492 and then to Korea by Japanese Army in order to kill Korean during the war, and the Aji was modified to Korean Kochu. In conclusion, in Korea our own Kochu was cultivated and used in the Korean native fermented foods such as kimchi and kochujang.

• Horticultural Science & Technology
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• v.34 no.6
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• pp.924-939
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• 2016

The great economic losses caused by Cucumber mosaic virus (CMV) infection of peppers has led to the development of genetically modified (GM) CMV-resistant peppers. We developed virus-resistant pepper plants using Agrobacterium tumefaciens -mediated transformation. The expressed recombinant protein was purified using nickel-nitrilotriacetic acid resin and immunoaffinity chromatography, and purity was assessed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Immunoblot analysis revealed the purified CMV coat protein (CMV-CP) had a molecular mass of 25 kDa. After in-gel digestion and desalting, the internal peptide fragments of CMV-CP were sequenced by matrix-assisted laser desorption/ionization-time of flight. Most GM pepper and Escherichia coli BL21 internal peptides had identical peptide sequences and contained 137 of 183 whole peptides in CMV-CP. A quantitative enzyme-linked immunosorbent assay was performed to detect CMV-resistant GM peppers. We also provide basic information about the expressed protein in GM peppers for further safety assessment. The contents of soluble protein and CMV-CP were measured in GM and control peppers cultivated in three different areas of Korea. Statistical significance in terms of cultivation areas, harvest times, generations, and plant tissue origin were determined based on a P value of 0.05. The highest amount of CMV-CP was detected at the seedling stage from plant grown in each region. T3 and T5 showed significantly different levels of CMV-CP from T4 in leaves in the whorl stage. No statistical differences were observed among GM peppers at different stages of maturity in any cultivation area. The results from this study contribute to the safety evaluation of newly designed CMV-resistant GM peppers and provide a standard against which to compare other virus-resistant GM peppers.

• Research in Plant Disease
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• v.15 no.3
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• pp.165-169
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• 2009

Twelve Cucumber mosaic virus (CMV) isolates were isolated from genetically modified (GM) and non-GM Capsicum annuum in two GM fields, Namyangju and Anseong, and their properties were investigated in this study. Coat protein (CP) gene of the CMV isolates were synthesized by RT-PCR using genus-specific primers which designed to amplify a DNA fragment of 950 bp. Purified cDNA fragments were cloned into the pGEMT easy vector for sequence determination. Nucleotide sequences (internal 657 bp) of CMV isolates were compared with Fny-CMV CP sequences and there were no significant collection site specific sequence similarities found. When predicted amino acid sequences (219 amino acids) were compared with Fny-CMV CP amino acids sequences, there were 96.8% to 97.3% similarities found from Namyangju collections and 95.9% to 96.8% similarities from Anseong collections. The phylogenetic analysis with nucleotide sequences showed definite differences in CMVs which have been isolated from the two regions.