DNA methylation at CpG sites, which is a epigenetic modification, is associated with gene expression without change of DNA sequences. During early mouse embryogenesis, dynamic changes of DNA methylation occur. In this study, DNA methylation patterns of porcine embryos produced in vivo and in vitro were examined at various developmental stages by the immunocytochemical staining method. Interestingly, active demethylation was not observed on the paternal pronucleus of porcine zygotes. However, differences were detected in the passive demethylation process between in vivo and in vitro embryos. There was no change in the DNA methylation state until the blastocyst stage of in vivo embryos, whereas partial demethylation was observed in several blastomeres from a 4 cell stage to a morula stage of in vitro embryos. The whole genome of inner cell mass (ICM) and trophectoderm (TE) cells in porcine blastocysts were evenly methylated without de novo methylation. Our findings demonstrate that genome-wide demethylation does not occur in pig embryos during preimplantation development unlike murine and bovine embryos. It indicates that the machinery regulating epigenetic reprogramming may be different between species.

사과원에서 사용되는 살비제 5종, 살충제 4종, 살균제 6종올 추천농도로 희석하여 점박이응애와 긴털이리응애에 대한 상대독성을 엽침지법으로 실험실에서 조사하였다. Dicofol은 점박이응애와 긴털이리응애에 대해 독성이 매우 강하였으며, 이들의 난에 대해서도 각각 46%와 40%의 致死效果가 있었다. Abamectin 용액에 침지된 긴털이리응애 암컷성충의 사망율은 무처리와 차이가 없었으나 점박이응애의 암컷성충은 24시간 이내에 모두 사망하였다. Tetradifon, Clofentezine, Hexythiazox등 3종의 살란성 살비제는 점박이 응애의 난에 대해서는 90%이상의 살란효과를 나타내었으나 긴털이리응애 난의 부화에는 아무런 영향이 없었다. Teflubenzuron, Triflumuron, Diflubenzuron, lmidacloprid 등 4종의 살충제와 Propineb, Fenarimol, Polyoxin-B.captan 등 3종의 살균제는 긴털이리응애 암컷성충에 대해 살비효과가 없었으며, Chlorothalonil, Nuarimol . Mancozeb, Folpet 등 3종의 살균제도 23% 이하의 낮은 살비효과를 나타내었다. 이상과 같이 긴털이리응애 난이나 암컷성충에 저독성을 나타내는 4종의 살충제와 4종의 살비제 및 6종의 살균제는 사과원 병해충 종합관리체계에서 긴털이리응애와 함께 이용될 수 있을 것으로 생각된다.

변색미(變色米) 발생(發生)에 관여(關與)하는 병원균(病原菌)과 해충을 분류동정(分類同定)한 결과(結果) 해충으로는 허리노린재과(科)에 속하는 Leptocorisa oratorius가 우점종(優占種)이었고 노린재과(科)인 Menida varipennis. Stollia ventralis 및 Nezara viridula 등이 관여(關與)하였으며 병원균(病原菌)으로는 Drechslera oryzae. Curvularia lunata, Trichoniella padwickii, Sarocladium oryzae, Alternaria tenuis 및 Fusarium solani 등이 관여(關與)하였다. 병원균(病原菌)과 해충의 복합발생시(複合發生時)에 변색미발생(變色米發生)이 더 심하였고 병원균(病原菌)만의 발생시(發生時)는 변색미(變色米) 발생(發生)에 주로 영향(影響)을 미쳤으며 노린재류만의 발생시(發生時)는 수량감수(收量減收)에 더 큰 영향(影響)을 주었다. 그리고 노린재류에 의한 벼 유숙기간(乳熟期間)의 흡즙(吸汁)은 병원균침입(病原菌侵入)을 조장(助長)하여 벼의 질적(質的) 변화(變化)와 양적(量的) 감소(滅少)에 크게 영향(影響)하였다.

Controllable transgenic expression systems in transgenic animal model are valuable to the development of therapeutic approaches in human medical fields. The aim of this study was to 1) produce a transgenic cloned dog using inducible tetracycline vector system, and 2) investigate whether the transgenic cloned dog could be induced the transgene expression using doxycycline (Doxy). Canine fetal fibroblasts were infected with retroviral vectors designed to express the enhanced green fluorescent protein (eGFP) gene under the control of tetracycline-inducible promoter. For somatic cell nuclear transfer (SCNT), nucleus of an in vivo matured oocyte was removed and an eGFP expressed cell cultured with 1 ㎍/㎖ of Doxy was injected. After electrical fusion and chemical activation, the reconstructed embryos were transferred to a recipient and pregnancy diagnosis was performed by ultrasonography. Experiment I evaluated the mean fluorescence intensity (MFI) of infected cells while the cells were cultured in the presence of 1 ㎍/㎖ of Doxy for 5 days, and then in the absence of Doxy for 7 days using fluorescence-activated cell sorter. Experiment II was designed to produce an eGFP controllable transgenic cloned dog via SCNT. For verification of transgenic dog, experiment III was performed Southern Blot analysis and observation in vivo regulation of eGFP expression in the cloned dog treated with 100 ㎎/㎏ of Doxy every 2 days for 2 weeks under ultraviolet light. In experiment IV, western blot was used to detect eGFP increase and decrease in skin tissues of transgenic dog under the presence or absence of Doxy. In the results of experiment I, the MFI for infected cells was rapidly increased to approximately 42.3 times after 3 day-treatment compared to pre-treatment and quickly decreased 3 days after ceasing the treatment. In experiment II, a total of 203 embryos were transferred to nine recipients and three pregnant delivered three pups (Tet-on eGFP 0, Tet-on eGFP 1, and Tet-on eGFP 2) by C-sec and Tet-on eGFP 2 among them is still alive. All cloned pups were genetically identical to the donor cell. Tet-on eGFP 2 showed an apparent in vivo eGFP expression on her body after Doxy administration in experiment III. The result of Sothern blotting showed that the transgene insertion was detected from the three cloned dogs and all organs of Tet-on eGFP 1. Experiment IV indicated that a robust eGFP expression in skin tissue of Tet-on eGFP 2 rapidly increased after Doxy treatment and gradually decreased to basal level on 9 weeks after ceasing the treatment. In conclusion, we report here for the first time an inducible transgenic system in canine species and it can stably induce the transgene expression at intended time. This study has demonstrated the capacity to generate transgenic model dog which could regulate the transgene and it would contribute to human medical research fields.