Herbicide-resistant transgenic rapeseed (TG rapeseed) was developed by inserting phosphinothricin acetyltransferase (PAT, bar), a modified gene from the soil bacterium Streptomyces hygroscopicus, into the genome of a conventional variety of rapeseed (Youngsan). The TG rapeseed used for the test was confirmed to express the PAT gene by polymerase chain reaction (PCR) and immunostrip. Feeding tests were conducted with Cyprinus carpio to evaluate the environmental risk of TG rapeseed, including the herbicide resistant gene. C. carpio was fed 100% ground rape suspension, TG rapeseed, or non-genetically modified (GM) counterpart rapeseed (Youngsan). As a result, the feeding test showed no significant differences in the cumulative immobility or abnormal response between C. carpio samples fed on TG rapeseed and non-GM counterpart rapeseed. The 48 h-LC50 values of the TG rapeseed and the non-GM counterpart rapeseed were 3,376 mg/L (95 % confidence limits: 3,169 - 3,596 mg/L) and 2,682 mg/L (95 % confidence limits: 2,267 – 3,123 mg/L), respectively. The rape NOEC (no observed effect concentration) value for C. carpio was suggested to be 625 mg/L. Based on these results, there was no significant difference in the toxicity for non-target organisms (C. carpio) between the TG rapeseed and non- GM counterparts.

GM작물은 세계적으로 재배면적이 지속적인 증가되고 있으며, 이로 인한 GM작물의 잠재적인 환경위해성에 대한 우려도 증가되고 있다. 현재까지 국내에서 GM작물의 상업적 재배는 되고 있진 않지만 GM작물의 안전성 평가를 위한 기술 개발은 크게 요구되고 있는 실정이다. 국내외적으로 다양한 GM작물의 유전자이동성 평가가 수행되어왔으나 화분 공급원인 GM 벼의 개화기 차이에 의한 유전자 이동성 연구는 부족한 실정이다. 본 연구에서는 벼의 개화기 차이에 따른 비타민A 강화 벼(PAC)로부터 모품종인 낙동벼와 중만생종인 일미벼, 조생종인 운광벼, 중생종인 대보벼, 통일형인 세계진미벼로의 화분 매개에 의한 유전자 이동성을 평가하였다. 비타민A 강화벼(PAC)의 도입 유전자를 검출하는 PCR 분석을 통해 유전자 이동성 유무를 최종적으로 검증하였다. 총 파종된 종자수에 대한 교잡율은 개화기가 일치하는 낙동벼에서는 0.0007%, 개화기가 차이가 있는 일미벼, 운광벼, 대보벼, 세계진미벼에서는 0%로 나타났으며, 모든 교잡개체들은 비타민A 강화벼(PAC)에 근접한 2m 내에서 발견되었다. 화분 매개에 의한 비타민A 강화벼(PAC)의 유전자 이동 특성은 기존에 연구된 결과들과 비슷한 결과를 보였으며, 벼 재배품종간의 개화기 차이가 화분에 의한 교잡을 결정하는데 중요한 요인으로 작용하였다. 이에 벼 경작지의 기상 조건과 벼 품종간의 개화시기 중복 여부 등이 GM벼에 의한 일반 재배품종 및 잡초성벼로의 유전자 이동 최소화 기술 개발과 안전관리 기준 작성에서 주요 영향 요소들로 고려해야 한다.

There are studies on the assessment of non-edible transgenic plants on soil microbial communities. In this research we evaluated the effect of virus-resistant trigonal cactus on soil microbial communities of the rhizosphere. Soil samples are collected and compared in genetically modified (GM) and non-GM trigonal cactus cultivation fields during vegetative growth period and post-harvest period. Biolog EcoplateTM was used to evaluate the functional diversity of soil microbial communities. There were no significant differences between the GM and non-GM soil samples collected during the vegetative growth period. However, we observed temporary difference in carbon substrate utilization. Principal component analysis showed that soil microbiota was influenced not by presence of GM or non-GM trigonal cacti, but rather by the cultivation period. Denaturing gradient gel electrophoresis fingerprinting revealed that virus-resistant trigonal cactus cultivation had insignificant effect on soil microbial communities including dominant rhizosphere bacteria, actinomycetes, and fungi. We found no clear evidence of GM trigonal cactus cultivation affecting the functional diversity of soil microbial communities.

The vitamin E enhanced transgenic soybean was developed by introducing a perilla γ- tocopherol methyltransferase gene (γ-TMT) under the control of pea vicilin promoter and a selection marker, phosphinothricin acetyltransferase (PAT) gene. With regard to the potential problems of safety, the non-target organism evaluation is required as an essential element for the environmental risk assessment of genetically modified (GM) crops. We studied the effects of the vitamin E enhanced transgenic soybean feeding on survival of Daphnia magna which is commonly used as a model organism in ecotoxicological studies. The Daphnia magna was fed on vitamin E enhanced transgenic soybean and non-genetically modified (non-GM) soybean (Willams 82) at 0, 1,000, 1,800, 3,240, 5,830, 10,500 and 20,000 mg/L concentrations, respectively. The GM soybean used for the test was confirmed to have the γ-TMT/PAT gene expression by the enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR) analysis. The results showed that there was no significant differences between cumulative immobilities of Daphnia magna fed on GM soybean and non-GM soybean. The 48hr-EC50 values showed no significant differences between GM soybean (2,416 mg/L) and non-GM soybean (2,408 mg/L). The results of this study suggested that there was no significant differences in toxicity for Daphnia magna between GM soybean and non-GM counterpart.

Genetically modified (GM) crops have been developed worldwide through the recombinant DNA technology and commercialized by global agricultural companies. Until now, GM crops have not been cultivated commercially in Korea. Commercialization of GM crops requires a compulsory assessment of environmental risk associated with the release of GM crops. This study was conducted to evaluate the frequency of pollen mediated gene flow from Bt transgenic rice (Agb0101) to japonica non-GM rice (Nakdongbyeo), indica non-GM rice (IR36), and weedy rice (R55). A total of 729,917, 596,318 and 230,635 seeds were collected from Nakdongbyeo, IR36, and R55, respectively, which were planted around Agb0101. Selection of the hybrids was determined by repeated spraying of herbicide and Cry1Ac1 immunostrip assay. Finally, the hybrids were confirmed by PCR analysis using specific primer. The hybrids were found in all non-GM rice and out-crossing ranged from 0.0005% at IR36 to 0.0027% at Nakdongbyeo. All of hybrids were located within 1.2 m distance from the Agb0101 rice plot. The meteorological elements including rainfall and temperature during rice flowering time were found to be important factors to determine rice out-crossing rate. Consideration should be taken for many factors like the meteorological elements of field and physiological condition of crop to set up the safety management guideline to prevention of GM crops gene flow.

This study was carried out to develop of environmental risk assessments and the biosafety guide for Vitamin E enhanced transgenic soybean at LMO (Living Modified Organism) isolation field. In LMO quarantine area of National Institute of Agricultural Sciences, insect species diversities and population densities on vitamin E enhanced transgenic soybean and non-GM soybeans (Willams 82 and Seoritae) were investigated. A total of 17,717 individuals of 77 species from 8 orders were collected in LMO isolation field. In three type soybeans field, total of 5,250 individuals in Vitamin E enhanced transgenic soybean, 5,510 individuals in Willams 82, and 6,957 individuals in Seoritae were collected, respectively. There was no difference between the population densities of insect pests, natural enemies and other insects on Vitamin E enhanced transgenic soybean and Willams 82, while natural enemies density on Seoritae was higher than on Vitamin E enhanced transgenic soybean, but insect pests density on Vitamin E enhanced transgenic soybean was higher. These results provided the insects diversity for risk assessment survey of Vitamin E enhanced transgenic soybean and suggested that the guideline could be useful to detect LMO crops.

The β-carotene biofortified transgenic rice was developed by transforming rice cv. Nakdongbyeo with phytoene synthase (Psy) and carotene desaturase (Crt I) genes isolated from Capsicum and Pantoea. The aim of this study was to perform molecular characterization of rice transformants of T5-T7 generation harboring Psy and Ctr I genes driven by endosperm specific globulin promoter for biosafety evaluation of β-carotene biofortified transgenic rice. The structure and sequence of T-DNA in the transformation vector and the insertion sites, flanking sequences and generational stability of inserted T-DNA in transgenic rice lines were analyzed. The transformation vector consisted of right border, MAR gene, carotenogenic genes unit, herbicide resistance selectable marker unit, MAR gene and left border in sequential order. T-DNA was introduced at the position of 30,363,938-30,363,973 bp of chromosome No. 2 by adaptor-ligation PCR. Stable integration of T-DNA and stable expression of bar gene was confirmed in T5 to T7 generations. It was also confirmed that the backbone DNA of transformation vector containing antibacterial gene was not present in the genome of β-carotene biofortified transgenic rice. HPLC analysis confirmed that carotenoids were consistently detected through T5-T7 generations.

Genetically modified (GM) crops have never been cultivated commercially in Korea, it is necessary for a thorough assessment of the risks associated with their environmental release. We determined the frequency of pollen mediated gene flow from disease resistant GM rice (OsCK1) to non-GM rice (Nagdongbyeo) and weedy rice (R55). A total of 449,711 or 164,604 seeds were collected from non-GM and weedy rice, respectively which were planted around OsCK1. Resistance of the hybrids was determined by repeated spraying of herbicide and DNA analysis using specific primer to confirm hybrids. Though non-GM rice and weedy rice have similar flowering time, the hybrids were found only in non-GM rice and out-crossing ranged from 0.018% at 0.3 m to 0.013% at 0.6 m. All of hybrids were located within 0.6 m distance from the GM rice plot in southerly direction. The meteorological factors including temperature and relative humidity during flowering time were found to be the most important factors for determining rice out-crossing. It should be considered many factors like the local weather condition and flowering time to set up the safety management policy to prevent pollen mediated gene flow between GM and conventional crop.

It is necessary to carry out a risk assessment to determine the consequences of releasing a particular plant species containing specific transgenes before transgenic plants can be grown under filed conditions. Gene flow from transgenic plants to wild closely related species has raised concern recently. Since transgenic crops were released in 1996, the global area of transgenic crops has been increasing rapidly. The transgene introgression from transgenic crops to their wild relatives is unavoidable in some species. Transgene introgression is of concern because the crop–wild plant hybrids might be conferred with a selection advantage to increase their performance, which could result in negative ecological consequences to natural ecosystems. The genus Brassica has 159 species, including a number of wild species that are of great importance to the economy. Most transgenic Brassica gene flow research has focused on the most successful cross between transgenic oilseed rape Brassica napus and its wild relatives Brassica rapa, a widely distributed weed in the farming system in Europe and America, since the hybridization can spontaneously happen and the generations can backcross to B. rapa easily in the wild conditions. In this study, we aimed to characterize transgene introgression, segregation, and expression in backcrossed generations between tramsgenic B. napus and B. rapa. These results will contribute to the environmental risk assessment and assist in biosafety management.

Internode elongation is an important agronomic trait in rice that is associated with lodging, yield, and flooding adaptation. We identified a novel rice mutant line showing shortened uppermost internode among the rice Ac/Ds insertional mutant population and named it shortened uppermost internode 4 (sui4). The phenotypes of F1 plants and F2 plants from the cross of sui4 with its original variety, Dongjin, indicated that the SUI4 gene shows incomplete dominance or semidominance. Because the Ds genotypes did not co-segregate with the sui4 phenotypes, we performed mapping of this gene with 273 F2 plants from a cross between sui4 and Milyang23. Primary mapping revealed that the SUI4 locus was located between the S07012 and S07015 markers on rice chromosome 7. Further fine mapping narrowed down the location of SUI4 to the 1.1-Mbp interval of RM1253-S07015. Using re-sequencing data of this mutant along with its original variety, Dongjin, and five other varieties, we found six sui4 specific SNPs occurred within the genic region of five genes in the fine-mapped interval. Among them, one SNP is in exon, while the other five SNPs are in intron. This SNP in exon occurred in the miR172 binding site of a gene encoding AP2 domain transcription factor, which seems to interrupt suppression of this gene activity by miR172. We isolated the genomic region of this gene from sui4 and transformed the wild type variety, Dongjin. The transgenic plants showed remarkably shortened internodes, which indicates that this AP2 domain transcription factor gene is the SUI4 gene.

In order to find new genetic sources of rice salt tolerance, we did screening with about 10,000 rice mutant lines created by Ac/Ds insertional mutagenesis. First, we raised rice seedlings with media soil on 0.7% NaCl solution and selected 71 putative salt tolerant lines and analyzed their Ds insertion sites. We tested their salt tolerance by growing seedlings on MS medium containing, 0 mM, 150 mM, and 250 mM NaCl. Also, their seedling salt tolerance were evaluated by growing on Yoshida nutritional solution containing 0.6% NaCl. Finally, we selected eight mutant lines showing increased seedling salt tolerance compared with wild type variety, Dongjin, repeatedly. We grow them in rice field and investigated their agronomic traits such as heading time, culm length, panicle length, and panicles per hill. Among them two lines which were named Salt10 and Salt23 and showed favorable agronomic characteristics were crossed with Dongjin for further genetic analysis and mapping the causative gene variation.

Over 7 individual rice (Oryza sativa L.) plants per a line were sowed and sampled by pooled sampling method for genomic DNA extraction. The 5,400 flanking sequence tags (FSTs) were analysed by adaptor PCR and direct sequencing. FST analysis showed that the intragenic FSTs, the intergenic FSTs, and the original insertional sequences including hot spot covered 48.1% (2,597), 25.6% (1,383), and 25% (1,350), respectively. The 2,597 intragenic FSTs were used for genotyping to determine whether they are heterozygous or homozygous, and 1,393 core lines were selected. Among them, 422 knockout genes were distributed on chromosome 3, while 56 - 157 intragenic FSTs scattered on other chromosomes. Among 1,393 FSTs, known genes such as transcription factor covered 59.4% (827), while unknown genes such as expressed protein covered 40.6% (566). RT-PCR indicated that some core lines had no expression or decreased expression level in their knockout genes. It means that core lines are very useful knockout lines for functional genomic studies.

1. Ds 삽입변이체 3,000계통으로부터 제초제 저항성 1,874계통을 선발하고 농업적 주요 특성으로서 출수일수, 간장, 수수, 수장, 엽장 등 5가지 형질에 대하여 조사한 바, 조사된 5가지 형질에 대하여 원품종인 동진벼에 비하여 매우 다양한 변이폭을 보여주었다. 2. 농업적 유용성과 관련된 수장이 길고, 조기출수, 수수가 많은 변이체 뿐만 아니라 형태학적 변이를 보이는 twin seedling, dwarf, early heading, strip albino, liguleless 등 변이체가 다수 발견됨으로서 육종적 이용 및 유전자 기능해석을 위한 유용한 집단으로서 유용성을 보여주었다. 3. 서던분석 결과, 벼 게놈상에서 Ds는 평균 2 copy로 전이되었으며 조직부위별로 GUS의 발현을 조사한 결과 잎, 뿌리 및 화기관등에서 약 3.9%가 발현되었다. 이 삽입변이체에서 나타난 다양한 변이형질의 주요 농업적 특성과 GUS 발현의 재현성을 위해 다음 세대의 전개를 통한 후대분석이 필요하다.