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RAPD 다형성 및 ABO 유전자형 분석을 통한 사상체질간 유전적 거리에 관한 연구
이휘철,조동욱,조중호,이창수 건국대학교 자연과학연구소 1999 建國自然科學硏究誌 Vol.10 No.2
본 연구는 사상체질인 태음인, 소음인 그리고 소양인간의 유전적 상관관계를 Random Amplified Polymorphic DNA(RAPD) 분석법 및 ABO 유전자형의 대립유전자의 빈도 분석을 통하여 실시하였다. 체질간 DNA 다형성을 검출할 수 있는 RAPD 분석용의 primer의 빠른 선별을 위해 체질별 혼합된 DNA 시료를 분석에 사용하였다. 일차적으로 200종류의 RAPD primer로부터 다형성을 보이는 7종류의 primer를 선별하였다. RAPD밴드로터 체질간 bandsharing(BS) 값은 0.68에서 0.71범위에 있었고, 유전적 거리는 BS 값에 의해 구했다. RAPD의 BS값에 의한 태음인과 소음인간 유전적 거리는 0.002이고, 이 두 체질과 소양인간에는 0.005로 분석되었다. 또한 체질별 ABO 유전형을 조사하여 그들의 대립유전자 빈도를 추정해 체질간 유전적 거리를 산출했다. 그 결과 태음인과 소음인사이에 유전적 거리는 0.014이고, 소양인과 나머지 두 체질간에는 0.032였다. 위와 같이 RAPD 분석법과 ABO 대립유전자의 빈도분석에 의한 체질간 상관관계는 두 방법 모두에서 태음인과 소음인이 소양인 체질보다 유전적으로 더 가깝게 나타났다. This study was carried out to be establish genetic understanding of Sasang constitutions of Teaumin, Soumin and Soyangin by Random Amplified Polymorphic DNA(RAPD) and ABO alleles frequencies analysis. We have applied RAPD analysis to pooled DNA sample as a means to achieve rapid screening of large numbers of primers for their capacity to reveal constitutions-specific polymorphisms. From initial 200 primers, 7 polymorphic primers between different constitutions were selected. The RAPD bandsharings(BS) values ranged from 0.68 to 0.71 for between three constitutions. The genetic distance between three constitutions was measured by BS values. Genetic distance by RADP analysis was 0.002 between Teaumin and Soumin, and 0.005 between Soyangin and the others. Three Sasang constitutions were investigated by the genotyping of ABO genotypes, and genetic distance was calculated from ABO allele frequencies in each constitution. As a result, genetic distance was 0.014 between Teaumin and Soumin, and 0.032 between Soyangin and the others. In conclusion, the genetic distance of Teaumin and Soumin was closer than that of Soyangin in the analysis of RAPD and ABO alleles frequencies.
Production of Recombinant Human Von Willebrand Factor in the Milk of Transgenic Pigs
LEE, Hyun-Gi,LEE, Hwi-Cheul,KIM, Sung Woo,LEE, Poongyeon,CHUNG, Hak-Jae,LEE, Yun-Keun,HAN, Joo-Hee,HWANG, In-Sul,YOO, Jong-Il,KIM, Yong-Kook,KIM, Hun-Taek,LEE, Hoon-Taek,CHANG, Won-Kyong,PARK, Jin-Ki Society for Reproduction and Development 2009 The Journal of reproduction and development Vol.55 No.5
<P>Von Willebrand factor (vWF), a large multimeric glycoprotein present in blood plasma, is a blood protein of the coagulation system. It is defective in von Willebrand disease and is involved in a large number of other diseases, including thrombotic thrombocytopenic purpura-hemolytic uremic syndrome and heyde's syndrome. We have developed a line of transgenic swine harboring recombinant human von Willebrand factor (rhvWF) cDNA through microinjection of fertilized one-cell pig zygotes. Expression of rhvWF in the mammary gland and secretion of rhvWF into the milk of the transgenic swine were confirmed by immunohistochemical and western blot analyses, respectively, and rhvWF proteins were detected in milk from all lactating founder females at concentrations that were 28- to 56-folds greater than that in circulating human plasma. The amino acid sequence of rhvWF protein in the transgenic pig milk matched that of vWF produced from human blood plasma. This study provides evidence that production of rhvWF from transgenic pig milk is a potentially valuable technology and can be used as a cost-effective alternative in clinical applications.</P>
Hwi-Cheul Lee,Gi-Sun Im,Hak Jae Chung,Poong-Yeon Lee,Jin-Ki Park,Won-Kyong Chang,Boh-Suk Yang,Keitaro Yamanouchi,Masugi Nishihara 사단법인 한국동물생명공학회 2006 Reproductive & developmental biology Vol.30 No.4
Our previous research has identified granulin (grn) and p130 genes as sex steroidinducible genes in the rat hypothalamus, which might be involved in sexual differentiation of the brain. Phthalate esters that are used as plasticizers and also found at low levels in foods such as dairy products are often mentioned as suspected endocrine disrupters. The purpose of the present study is to elucidate whether perinatal exposure to dinbutyl phthalate (DBP), diisononyl phthalate (DINP) and di2ethylhexyl adipate (DEHA) affects hypothalamic sex steroidinducible genes. The present study assessed the effects of perinatal exposure to DBP, DINP and DEHA on sex steroid hormones levels and hypothalamic grn and p130 mRNA expressions at postnatal day (PND) 3 and 7. Pregnant rats were fed a soyfree diet containing 20, 200, 2,000 and 10,000 ppm of DBP, 40, 400, 4,000 and 20,000 ppm of DINP, or 480, 2,400 and 12,000 ppm of DEHA from gestational day (GD) 15 to GD 3 or 7. At PND 3 and 7, perinatal exposure to these chemicals did not substantially affect serum concentrations of testosterone and estradiol. At PND 3, the expression of grn mRNA levels in males was decreased by DEHA, and that of p130 was decreased by DBP, DINP and DEHA, though the effects were not dosedependent. At PND 7, the expression of grn gene in female pups was increased by higher doses of DBP and all the doses, except for 4,000 ppm, of DINP, while that in male pups decreased by 480 and 12,000 ppm of DEHA. Hypothalamic expression of p130 mRNA in males was increased by lower doses of DBP and all the doses of DINP, whereas that of females was decreased by 480 and 2,400 ppm of DEHA. These results suggest that these chemicals may affect the expression of grn and p130 genes by directly acting on the hypothalamus, thus leading to inappropriate expression of these genes.
Lee, Hwi-Cheul,Yang, Byoung-Chul,Ko, Yeoung-Gyu,Kim, Dong-Hoon,Park, Jin-Ki,Yang, Boh-Suk,Seong, Hwan-Hoo,Yamanouchi, Keitaro,Nishihara, Masugi The Korean Society of Animal Reproduction 2006 Reproductive & developmental biology Vol.30 No.4
Di-n-butyl phthalate (DBP), diisononyl phthalate (DINP) and di-(2-ethylhexyl) adipate (DEHA) are ubiquitously distributed chemicals that are widely used as plasticizers and also found at low levels in foods. The aims of this study were to determine whether perinatal exposure to DBP, DINP and DEHA could alter normal patterns of neonatal development. Dams were provided with pulverized soy-free diet containing 20, 200, 2,000 and 10,000 ppm of DBP, 40, 400, 4.000 and 20,000 ppm of DINP, or 480, 2,400 and 12,000 ppm of DEHA from gestational day 15 to postnatal day 21. Exposure to the high doses of DBP, DINP and DEHA during gestational period significantly decreased food consumption and body weight gain of dams. These chemicals reduced neonatal body weight as well as that of the after maturation. Also, exposure to DINP of all the doses used and the higher doses (2,400 and 12,000 ppm) of DEHA decreased AGD at PND 1 in male neonates, though that to DBP did not affect AGD in males. In female neonates, an increase in AGD was observed in DBP- and DINP-exposed animals at the highest doses. Moreover, these chemicals affected survival rate of pups at PND 5, and delayed onset of eye opening in all chemica1-exposed groups at PND 17. These results suggest that perinatal exposure to these chemicals may affect the normal development and/or growth of offspring.
Lee, Hwi-Cheul,Im, Gi-Sun,Chung, Hak-Jae,Lee, Poong-Yeon,Park, Jin-Ki,Chang, Won-Kyong,Yang, Boh-Suk,Yamanouchi, Keitaro,Nishihara, Masugi The Korean Society of Animal Reproduction 2006 Reproductive & developmental biology Vol.30 No.4
Our previous research has identified granulin (grn) and p130 genes as sex steroid-inducible genes in the rat hypothalamus, which might be involved in sexual differentiation of the brain. Phthalate esters that are used as plasticizers and also found at low levels in foods such as dairy products are often mentioned as suspected endocrine disrupters. The purpose of the present study is to elucidate whether perinatal exposure to di-n-butyl phthalate (DBP), diisononyl phthalate (DINP) and di-2-ethylhexyl adipate (DEHA) affects hypothalamic sex steroid-inducible genes. The present study assessed the effects of perinatal exposure to DBP, DINP and DEHA on sex steroid hormones levels and hypothalamic gm and p130 mRNA expressions at postnatal day (PND) 3 and 7. Pregnant rats were fed a soy-free diet containing 20, 200, 2,000 and 10,000 ppm of DBP, 40, 400, 4,000 and 20,000 ppm of DINP, or 480, 2,400 and 12,000 ppm of DEHA from gestational day (GD) 15 to GD 3 or 7. At PND 3 and 7, perinatal exposure to these chemicals did not substantially affect serum concentrations of testosterone and estradiol. At PND 3, the expression of grn mRNA levels in males was decreased by DEHA, and that of p130 was decreased by DBP, DINP and DEHA, though the effects were not dose-dependent. At PND 7, the expression of gm gene in female pups was increased by higher doses of DBP and all the doses, except for 4,000 ppm, of DINP, while that in male pups decreased by 480 and 12,000 ppm of DEHA. Hypothalamic expression of p130 mRNA in males was increased by lower doses of DBP and all the doses of DINP, whereas that of females was decreased by 480 and 2,400 ppm of DEHA. These results suggest that these chemicals may affect the expression of gm and p130 genes by directly acting on the hypothalamus, thus leading to inappropriate expression of these genes.
Hwi-Cheul Lee,Byoung-Chul Yang,Yeoung-Gyu Ko,김동훈,Jin-Ki Park,Boh-Suk Yang,Hwan-Hoo Seong,Keitaro Yamanouchi,Masugi Nishihara 사단법인 한국동물생명공학회 2006 Reproductive & developmental biology Vol.30 No.4
Di-n-butyl phthalate (DBP), diisononyl phthalate (DINP) and di-(2-ethylhexyl) adipate (DEHA) are ubiquitously distributed chemicals that are widely used as plasticizers and also found at low levels in foods. The aims of this study were to determine whether perinatal exposure to DBP, DINP and DEHA could alter normal patterns of neonatal development. Dams were provided with pulverized soy-free diet containing 20, 200, 2,000 and 10,000 ppm of DBP, 40, 400, 4,000 and 20,000 ppm of DINP, or 480, 2,400 and 12,000 ppm of DEHA from gestational day 15 to postnatal day 21. Exposure to the high doses of DBP, DINP and DEHA during gestational period significantly decreased food consumption and body weight gain of dams. These chemicals reduced neonatal body weight as well as that of the after maturation. Also, exposure to DINP of all the doses used and the higher doses (2,400 and 12,000 ppm) of DEHA decreased AGD at PND 1 in male neonates, though that to DBP did not affect AGD in males. In female neonates, an increase in AGD was observed in DBP- and DINP-exposed animals at the highest doses. Moreover, these chemicals affected survival rate of pups at PND 5, and delayed onset of eye opening in all chemical-exposed groups at PND 17. These results suggest that perinatal exposure to these chemicals may affect the normal development and / or growth of offspring.
Study on the Reproductive Function in Transgenic Pig Harboring Human Erythropoietin (hEPO) Gene
Lee, Hyun-Gi,Lee, Hwi-Cheul,Chung, Hak-Jae,Hwang, In-Sul,Choi, Myoung-Seob,Byun, Sung-June,Lee, Seung-Hoon,Kim, Min-Ji,Woo, Jae-Seok,Chang, Won-Kyong,Lee, Poong-Yeon,Lee, Hoon-Taek,Park, Jin-Ki The Korean Society of Animal Reproduction 2008 Reproductive & developmental biology Vol.32 No.2
Our previous study showed that transgenic (TG) pigs harboring human EPO (hEPO) gene have been shown to have reproductive disorders, including low pregnancy rates, irregular estrus cycle and low little size. To investigate these reasons, we assessed estrus behavior (standing response) and plasma $17{\beta}$-estradiol ($E_2$) level, which partly reflect reproductive function, during the estrus cycles after synchronization and superovulation by hormone treatments. Then, we analysed blood composition and expression of hEPO gene in TG pigs. Pigs were injected with PG600. After 10 days, pigs were fed with Regumate porcine for 6 days. Blood samples were collected from jugular vein. Analysis of blood composition and $E_2$ level were measured by Hemavet 950 and $E_2$ ELISA kit, respectively. And, the expression of hEPO gene in reproductive organs was quantitated by real-time RT-PCR. The percentage of estrus behavior in TG was significantly decreased. Hematocrit (HCT), hemoglobin (Hb) concentration and red blood cell (RBC) number were significantly higher in TG than wild type (WT). On the other hand, high expression of hEPO gene in TG was observed in the mammary gland as well as in the uterus. Moreover, plasma $E_2$ level was significantly higher in TG than WT. These results suggest that nonspecific expression of hEPO gene in the other organs of TG may affect blood composition and plasma $E_2$ level, thereby causing reproductive disorders.
LEE, Hwi-Cheul,YAMANOUCHI, Keitaro,NISHIHARA, Masugi 家畜繁殖硏究所 2006 Journal of Reproduction and Development Vol.52 No.3
<P>Our previous research has identified the granulin (grn) and p130 genes as sex steroid-regulated genes in the neonatal rat hypothalamus that might be involved in sexual differentiation of the brain. Since phthalate/adipate esters such as di-n-butyl phthalate (DBP), diisononyl phthalate (DINP), and di-2-ethylhexyl adipate (DEHA) are suspected to interfere with the endocrine system as environmental endocrine disruptors having estrogenic or antiandrogenic properties, these chemicals may affect sexual differentiation of the brain. The present study assessed the effects of perinatal exposure to DBP, DINP, and DEHA on grn and p130 mRNA expressions in the hypothalamus on postnatal day (PND) 7 and sexual behaviors after maturation in rats. Maternal rats were given a phytoestrogen-free diet containing different doses of DBP (20, 200, 2,000, and 10,000 ppm), DINP (40, 400, 4,000, and 20,000 ppm) and DEHA (480, 2,400, and 12,000 ppm) from gestational day 15 to the day of weaning (PND 21). DBP and DINP exposure during the perinatal period resulted in an increase in hypothalamic grn and p130 mRNA levels in females and males, respectively, but DEHA exposure decreased expression levels of grn in males and p130 in females, although the effects were not dose-dependent. After maturation, male rats that were exposed to several doses of DBP, DINP, and DEHA displayed decreased copulatory behavior. The lordosis quotient was decreased in females perinatally exposed to DBP, DINP, and DEHA at all the doses used. On the other hand, serum levels of LH and FSH in both sexes and the estrous cycles in females were not affected by the treatments. These results suggest that inappropriate expression of grn and/or p130 genes in the brains of male and female neonatal rats by perinatal exposure to these chemicals may exert permanent effects on the hypothalamus, thereby decreasing sexual behavior after maturation.</P>
Comparison of Identification Methods for Transgene and the Production of GFP dlabetic Transgenic Pig
Hwi-Cheul Lee,Hyun-Gi Lee,Hee Kyoung Chung,Seunghoon Lee,Lee, Min-Ji Kim,Sung-June Byun,Jae-Seok Woo,Eun-Mi Ko,Won-Kyoung Chang,Jin-Ki Park 한국동물번식학회 2008 Reproductive & Developmental Biology(Supplement) Vol.32 No.2s