• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2001년도 춘계학술발표대회
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• pp.51-51
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• 2001

Cocaine and amphetamine-regulated transcript (CART) is a satiety factor that is regulated by leptin. It was reported that the mice intracerebroventricularly injected with CART showed behavioral changes resembled with the typical behavioral alterations found in the mice carrying disorders in the brain serotonergic (5-HT) system. Hence, this study was conducted to find out the relationships between CART and 5-HT. We first examined the mRNA levels of CART after the injections of para-chlorophenylalanine (pCPA, 300 mg/kg i.p., single injection or daily for three consecutive days) in the rat brains by in situ hybridization using the mouse CART cDNA probe cloned in our laboratory. Systemic administrations of pCPA, a potent inhibitor of tryptophan hydroxylase, the rate limiting enzyme of 5-HT biosynthesis, acutely depletes the brain 5-HT transporter (5-HTT) in the dorsal raphe nucleus (DRN), which reuptakes terminal 5-HT. Results indicated that the mRNA level of CART significantly decreased in the arcuate nucleus, paraventricular nucleus, and lateral hypothalamic nucleus by three days of daily injection with pCPA with no noticeable change detected 24 hrs after the single injection. The message levels of 5-HTT in DRN decreased in both single and three days of injections. Secondly, to investigate whether CART affect to 5-HT, mouse genomic CART gene, which is consist of 3 exons and 2 introns and mouse neurofilament light (NF-L) chain promoter were cloned. Then, we constructed neuron specific expression vector, which was transfected into HeLa cell using lipid-mediated transfection system. Expression of GFP and CART linked to NF-L-chain promoter in the transfected HeLa cell were detected by using fluorescent microscope and RT-PCR. These results confirmed normal expression of DNA constructs in vitro. Then, to increase brain specific expression of CART in vivo transgenic mice carrying CART gene controlled the deleted NF-L-chain promoter were generated by the DNA microinjection into pronuclei of fertilized embryos. Transgenic mice were detected by Southern blot. Further study is necessary to examine CART expression and 5-HTT in these transgenic mice. Therefore, these results suggest that there maybe a positive molecular correlation between CART and 5-HT in responding to the stimuli.

• Asian-Australasian Journal of Animal Sciences
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• 제20권5호
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• pp.615-621
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• 2007

AMP-activated protein kinase alpha 2 (PRKAA2) plays a key role in regulation of fatty acid and cholesterol metabolism. This study investigated the porcine PRKAA2 gene as a positional candidate for intramuscular fat and backfat thickness traits in pig chromosome 6. A partial fragment of the porcine PRKAA2 gene, amplified by PCR, contained a putative intron 3 including a part of exon 3 and 4, comparable with that of human PRKAA2 gene. Within the fragment, several single nucleotide polymorphisms were identified using multiple sequence alignments. Of these, TaqI restriction enzyme polymorphism was used for genotyping various pig breeds including Korean reference family. Using linkage and physical mapping, the porcine PRKAA2 gene was mapped in the region between microsatellite markers SW1881 and SW1680 on chromosome 6. Allele frequencies were quite different among pig breeds. The full length cDNA of the porcine PRKAA2 (2,145 bp) obtained by RACE containing 1,656 bp open reading frame of deduced 552 amino acids, had sequence identities with PRKAA2 of human (98.2%), rat (97.8%), and mouse (97.5%). These results suggested that the porcine PRKAA2 is a positional candidate gene for fat deposition trait at near telomeric region of the long arm of SSC 6.

• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 1998년도 제4차 학술발표대회 및 정기총회
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• pp.61-62
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• 1998

The effects of rearing temperature on the sex determination of zebrafish were examined to address the questions, "Is it possible to control phenotypic sex artificially; and what genes may be involved in that process\ulcorner". At higher temperature(${32-34}^{\circ}C vs 28.5^{\circ}$C), a dramatic shift of the sex ratio(female 1 : male 9) was found. with DD-PCR cloning, several candidate genes were cloned and analyzed. Further analysis should help to understand sex-determining mechanisms involved in the artificial induction.induction.

• 한국임상수의학회:학술대회논문집
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• 한국임상수의학회 2007년도 추계국제학술대회 및 컨퍼런스
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• pp.566-566
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• 2007

• 한국가금학회지
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• 제46권1호
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• pp.17-24
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• 2019

본 연구는 오브알부민 프로모터의 최적 크기를 결정하기 위해 수행하였고, 이를 위해 2.8, 5.5, 그리고 Mut-4.4 kb의 오브알부민 프로모터를 리포터 벡터에 클로닝하여 다양한 세포들에서 프로모터의 활성을 검증하였다. Mut-4.4kb-pOV는 2.8와 5.5kb-pOV에 비해 HeLa, MES-SA, 그리고 LMH/2A에서 높은 수준의 프로모터 활성을 유도하였으나, cEF 세포에서는 낮은 활성을 보였다. 한편, Mut-4.4kb-pOV/pGL4.11 벡터가 도입된 HeLa, MES-SA, LMH/2A, 그리고 cEF 세포에서 에스트로겐 처리에 의한 반응성을 검증한 결과, cEF 세포를 제외한 나머지 세포들은 에스트로겐을 처리했을 때 리포터 유전자의 발현량이 증가하였다. 또한 LMH/2A 세포에 500 nM 에스트로겐 처리 결과, Mut-4.4kb-pOV는 에스트로겐 처리 후에도 2.8와 5.5kb-pOV에 비해 높은 수준의 프로모터 활성을 유도하였다. 더불어 ERE 영역이 없는 2.8kb-pOV는 LMH/2A 세포에서 500 nM 에스트로겐 처리 후 프로모터의 활성에 효과가 없었으나, ERE 영역을 포함하는 5.5 그리고 Mut-4.4kb-pOV는 에스트로겐 처리에 의해 프로모터의 활성이 증가한 결과를 보였다. 이상의 결과는 Mut-4.4kb-pOV가 형질전환 암탉을 생산하기 위한 재조합바이러스 벡터의 프로모터 영역으로 2.8와 5.5kb-pOV에 비해 보다 효율적인 크기임을 보여주는 결과이다.

• Asian-Australasian Journal of Animal Sciences
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• 제22권5호
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• pp.712-720
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• 2009

The goal of this study was to elucidate the expression and segmental distribution of the glomerular cationic amino acid metabolism transporter-2 (CAT-2) and thus to improve our understanding of porcine cationic amino acid transporters and amino acid absorption. Porcine CAT-2 was cloned, sequenced and characterized. The predicted amino acid sequence of porcine CAT-2 shared 86.1% and 92.1% identity with human and mouse CAT-2A, respectively. The tissue distribution patterns and ontogenic changes of CAT-2 mRNAs were determined by real-time Q-PCR. The results showed that porcine CAT-2 was highly expressed in the heart and intestinal tract (duodenum, ileum and jejunum). In addition, the mRNA of CAT-2 was found in liver, lung, kidney, brain and muscle. Within the intestinal tract, CAT-2 mRNA was most abundant in the ileum and rarely expressed in the duodenum. In the duodenum, the levels of CAT-2 mRNA reached their peak on day 7 (p<0.05) while in the jejunum, levels were low on day 1 and 7 and increased rapidly after day 26 before peaking on days 30 and 60 (p<0.05). The levels then dramatically decreased by day 90 (p<0.05). In the ileum, levels achieved their maximum on day 30 and then decreased significantly on day 60 (p<0.05).

• Asian-Australasian Journal of Animal Sciences
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• 제23권9호
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• pp.1159-1165
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• 2010

Glucose is the main energy source for mammalian cells and its absorption is co-mediated by two different families of glucose transporters, sodium/glucose co-transporters (SGLTs) and facilitative glucose transporters (GLUTs). Here, we report the cloning and tissue distribution of porcine GLUT2. The GLUT2 was cloned by RACE and its cDNA was 2,051 bp long (GenBank accession no. EF140874). An AAATAA consensus sequence at nucleotide positions 1936-1941 was located upstream of the poly $(A)^+$ tail. Open reading frame analysis suggested that porcine GLUT2 contained 524 amino acids, with molecular weight of 57 kDa. The amino acid sequence of porcine GLUT2 was 87% and 79.4% identical with human and mouse GLUT2, respectively. GLUT2 mRNA was detected at highest level in porcine liver, at moderate levels in the small intestine and kidney, and at low levels in the brain, lung, muscle and heart. In the small intestine, the highest level was in the jejunum. In conclusion, the mRNA expression of GLUT2 was not only differentially regulated by age, but also differentially distributed along the small intestine of piglets, which may be related to availability of different intestinal luminal substrate concentrations resulting from different food sources and digestibility.

• 한국가축번식학회지
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• 제24권1호
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• pp.49-57
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• 2000

본 연구는 소 태아섬유아세포를 이용하여 핵이식 후 세포의 휴면처리, 세포의 passage 수 및 세포의 기원이 복제란의 발육에 미치는 영향을 검토하였다.3.57개 월령 한우 수컷 태아의 피부 및 간 조직에서 세포를 채취하여 체외에서 4∼6 회 계대배양 후 동결하였다가 핵이식에 사용하였다. 세포는 핵이식 전에 혈청기아처리를 하였으며, 대조구로는 활발히 분열 중의 무처리 세포를 사용하였다. Donor 세포는 미수정란의 탈핵세포질에 이식 후 전기융합 및 활성화를 실시하였고, 재구축배는 7∼9 일간 체외배양하여 발육농을 검토하였다. 배반포로 발육된 일부 복제란은 발정 7∼8 일째의 수란우에 이식하였다. 복제란의 배반포 발육율은 혈청기아 처리구 (25.3%)가 무처리구 (15.9%)에 비하여 유의적으로 높았으나 (P＜0.05), 세포의 passage 수 (4∼6회) 에 관계없이 23.1∼25.0%로 나타났고, 피부 및 간유래 복제란의 배반포 발육율도 23.8∼25.2% 로 두 조직세포 간에 차이가 없었다. 복제란의 이식 후 60일 및 120일에 수란우의 34.4% 및 15.6%가 각각 임신이 확인되었으며, 초자화동결된 태아 피부세포 복제란으로부터 1두의 수컷 송아지가 생산되었다. 본 연구의 결과는 복제란의 체외발육능이 세포의 휴면처리에 의하여 향상되었으나, 세포의 passage 수 (4∼6 회) 및 세포의 두 기원 (피부 및 간) 에 의해서는 영향을 받지 않으며, 태아 피부세포 유래 복제란으로부터 산자가 생산될 수 있음을 확증한다.

• BMB Reports
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• 제39권1호
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• pp.22-25
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• 2006

The gene encoding MPB83 from Mycobacterium bovis Vallee111 chromosomal DNA was amplified by using polymerase chain reaction (PCR) technique, and the PCR product was approximately 600bp DNA segment. Using T-A cloning technique, the PCR product was cloned into pGEM-T vector and the cloning plasmid pGEM-T-83 was constructed successfully. pGEM-T-83 and pET28a(+) were digested by BamHI and EcoRI double enzymes. The purified MPB83 gene was subcloned into the expression vector pET28a(+), and the prokaryotic expression vector pET28a-83 was constructed. Plasmid containing pET28a-83 was transformed into competence Escherichia coli BL21 (DE3). The bacterium was induced by isopropyl-$\beta$-D-thiogalactopyranoside (IPTG) and its lysates were loaded directly onto sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE), approximately 26 kDa exogenous protein was observed on the SDS-PAGE. The protein was analyzed using Western-blotting. The results indicated that the protein was of antigenic activity of M. bovis. The results were expected to lay foundation for further studies on the subunit vaccine and DNA vaccine of MPB83 gene in their prevention against bovine tuberculosis.

• Journal of Microbiology and Biotechnology
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• 제25권11호
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• pp.1960-1965
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• 2015

Rabbit hemorrhagic disease virus (RHDV) is highly contagious and often causes fatal disease that affects both wild and domestic rabbits of the species Oryctolagus cuniculus. A highly pathogenic RHDV variant (RHDVa) has been circulation in the Korean rabbit population since 2007 and has a devastating effect on the rabbit industry in Korea. A highly pathogenic RHDVa was isolated from naturally infected rabbits, and the gene encoding the VP60 protein was cloned into a baculovirus transfer vector and expressed in insect cells. The hemagglutination titer of the Sf-9 cell lysate infected with recombinant VP60 baculovirus was 131,072 units/50 μl and of the supernatant 4,096 units/50 μl. Guinea pigs immunized twice intramuscularly with a trial inactivated RHDVa vaccine containing recombinant VP60 contained 2,152 hemagglutination inhibition (HI) geometric mean titers. The 8-week-old white rabbits inoculated with one vaccine dose were challenged with a lethal RHDVa 21 days later and showed 100% survival rates. The recombinant VP60 protein expressed in a baculovirus system induced high HI titers in guinea pigs and rendered complete protection, which led to the development of a novel inactivated RHDVa vaccine.