• Proceedings of the Korean Society of Developmental Biology Conference
• /
• 2003.10a
• /
• pp.101-101
• /
• 2003

Main strategy for a treatment of Parkinson's disease (PD), due to a progressive degeneration of dopaminergic neurons, is a pharmaceutical supplement of dopamine derivatives or ceil replacement therapy. Both of these protocols have pros and cons; former exhibiting a dramatic relief but causing a severe side effects on long-term prescription and latter also having a proven effectiveness but having availability and ethical problems Embryonic stem (ES) cells have several characteristics suitable for this purpose. To investigate a possibility of using ES cells as a carrier of therapeutic gene(s), human ES (hES, MB03) cells were transfected with cDNAs coding for tyrosine hydroxylase (TH) in pcDNA3.1 (+) and the transfectants were selected using neomycin (250 $\mu /ml$). Expression of TH being confirmed, two of the positive clone (MBTH2 & 8) were second transfected with GTP cyclohydrolase 1 (GTPCH 1) in pcDNA3.1 (+)-hyg followed by selection with hygromycin-B (150 $\mu /ml$) and RT-PCR confirmation. By immune-cytochemistry, these genetically modified but undifferentiated dual drug-resistant cells were found to express few of the neuronal markers, such as NF200, $\beta$-tubulin, and MAP2 as well as astroglial marker GFAP. This results suggest that over-production of BH4 by ectopically expressed GTPCH I may be involved in the induction of those markers. Transplantation of the cells into striatum of 6-OHDA- denervated PD animal model relieved symptomatic rotational behaviors of the animals. Immunohistochemical analyses showed the presence of human cells within the striatum of the recipients. These results suggest a possibility of using hES cells as a carrier of therapeutic gene(s).

• Journal of Animal Science and Technology
• /
• v.51 no.3
• /
• pp.193-200
• /
• 2009

The pork from black-coated pigs is famous among-consumers for better eating quality. The loci affecting black coat color was identified in pig chromosome 6 in which several genetic effects on pork quality have been reported. The melanocortin 1 receptor (MC1R) gene is a major gene which plays a key role in regulation of eumelanin (black/brown) and phaeomelanin (red/yellow). In this study, the MC1R gene polymorphism was analyzed for pig breed determination and genetic association with pork quality traits. MC1R Ala243Thr variation was analyzed to determine a specific genotype for four commercial pig breeds (Landrace, Yorkshire, Berkshire and, Duroc) and a Korean native pigs (KNP). Then we developed original KNP-specific DNA markers to determine the pork from black-coated pigs using MC1R DNA sequences. The total length of the MC1R coding sequence ranged 1451bp in KNP. KNP had the 0201 allele pertaining to $E^{D1}$ but some of the KNP had the $E^P$ allele, probably reflecting the geneticintrogression of $E^P$ allele into KNP. Furthermore, a relationship between Leu102Pro single nucleotide polymorphism (SNP) genotype and pork quality phenotype were analyzed in F2 reciprocal-crossbred population between KNP and Yorkshire. Association analysis indicated that the allele of the MC1R gene has no effect on pork quality. These results suggest that black coat-color is not directly associated with preferred pork quality, but the black-coat color pig breed may have other genetic components for superior pork quality.

• Korean Journal of Biological Psychiatry
• /
• v.2 no.1
• /
• pp.57-62
• /
• 1995

Objects : Clozapine, prototype of the atypical neuroleptics, was known to have unique antipsychotic effect with a few extrapyramidal effects. While most typical antipsychotic agents mainly block $D_2$ receptors, clozapine has higher affinity for dopamine $D_4$ receptor than for $D_2$ receptor. Many researchers have tried to find out the relationship between schizophrenia and the abnormality of the genes coding dopamine receptors. But no consistent findings were reported. Recently, dopamine $D_4$ receptor was fully sequenced, and the alleles of dopamine $D_4$ receptor gene was found in unusual form on the 48th base pair. Our study was performed to identify the distribution of the dopamine $D_4$ receptor alleles of schizophrenics and normal controls, and whether any difference between the dopamine $D_4$ receptor alleles of schizophrenics and that of normal controls exists. Methods : DNA was extracted from the blood of schizophrenic patients(N=60) and normal controls(N=60). Part of the dopamine $D_4$ receptor gene was amplified by PCR, and amplified DNA was electrophoresed. Authors compared the distribution of the alleles of dopamine $D_4$ receptor gene of normal controls and that of schizophrenic patients. Results : Six kinds of alleles of $D_4$ receptor were observed both groups. The fourth repeat form of alleles was the most common in both schizophrenic patients(75.8%) and normal controls(70.3%), so there was not significant difference between two groups. Conclusion : The Difference in the distribution of the dopamine $D_4$ receptor gene alleles is not thought to be responsible for the pathophysiology of the schizophrenia. However, the difference in the expression of the dopamine $D_4$ receptor gene between normal and schizophrenia is left to be scrutinized.

• Journal of Plant Biotechnology
• /
• v.33 no.1
• /
• pp.27-32
• /
• 2006

To clarify the roles of bromelain in plants, we isolated BL1 gene encoding bromelain from pineapple stem tissues and sequenced. The full length cDNA is 933 bp and encodes a polypeptide of 311 amino acid residues. The cDNA is most similar 94% at the amino acid level to bromelain previously isolated from pineapple (BAA21929). Explants of Lactuca sativa were co-cultivated with Agrobacterium tume-faciences LBA 4404 strains containing nptII and BL1 gene for transformation. Through initial selection of regenerated explants by culturing on a kanamycin and carbenicillin containing MS medium, multiple shoots were obtained after 2 months of culture. For a complementary step of selection, putative transgenic shoots were transferred to 1/2 Ms basal medium supplemented with 100 mg/L kanamycin and 500 mg/L carbenicillin. The selected shoots were obtained T1 generation seeds with emasculation, and tested with PCR analysis using 35S promoter and BL1 specific primers whether BL1 gene was introduced to genome of the plants. These results confirmed that produced the specific PCR bands in the putative transgenic lines. Additionally the Northern blot and endo protease activity showed that transcripts of BL1 gene were detected in transgenic lines. Theses results suggest that BL1 gene be successfully integrated and transcripted in the transgenic lettuce plants.

• Microbiology and Biotechnology Letters
• /
• v.29 no.1
• /
• pp.18-24
• /
• 2001

Expression of the baculovirus major envelope glycoprotein gene(gp64) is regulated by transcription from botha early and late promoters. To develop a transient expression vector under the control of gp64 gene promoter, the gp64 gene of Bombyx mori nucleopolyhedrovirus-K1(BmNPV-K1) was characterized. The gp64 gene was local-ized at EcoR I-Pst I 7.38-kb fragment of the BmNPV-K1 genome. The EcorR 1-Pst I 7.38-kb fragment was cloned and the nucleotide sequence of 2,277 bases including the coding region of gp64 gene was determined. Based on these results, transient expression vector using gp64 gene promoter was constructed and named as pBm64. E.coli lacZ gene was introduced onto pBm64 as a reporter gene and expressed transiently in B. mori 5(Bm 5) cells. The expression vector transfected into the cells was maintained stably for 1 to 5 days. In order to confirm the expression of the reporter gene by gp64 promoter, recombinant virus was constructed. The recombinant virus has two independent transcription units in opposite orientations with two promoters; gp64 and polyhedrin gene promoters each initiating transcription of $\beta$-galactosidase and polyhedrin, respectively. Polyhedra formation and expression of $\beta$-galactosidase in Bm5 cells infected with the recombinant virus were observed with phase contrast microscope and in situ staining.

• Journal of Animal Science and Technology
• /
• v.45 no.2
• /
• pp.169-182
• /
• 2003

To understand the structure and regulation of bovine ADRP (Adipocyte Differentiation Related Protein) gene, we have isolated the genomic clone of bovine ADRP and determined its sequence. A genomic Southern blot analysis confirmed that ADRP gene is present as a single copy in bovine genome and the ADRP gene spans 12 kb. Bovine ADRP genomic clone, HwADRPg-1, had 8 exons and 7 introns, and all splicing sites conformed to the GT/AG rule with the exon-intron boundaries located exactly. Analysis of the upstream 649 bp of the sequence of HwADRPg-1 showed that it does not contain any canonical TATAA boxes; however Sp1 binding sites and CAAT boxes are found. The promoter contained potential binding sites for AP-1, AP-2 and several putative transcription factor binding sites. The 5'-flanking region of HwADRPg-1 contained muscle specific transcription activator Myo G and C/EBP (CCAAT/ enhancer binding protein) recognizing site. These results suppose that the Myo G transcription activator regulate the transcription of bovine ADRP gene in muscular tissue and its transcriptional activity was triggered by degree of muscular development. Our results provide the necessary analysis for other flanking sequences are needed in addition to the proximal cis elements of this promoter to confer adipocyte differentiation-dependent or growth-dependent transcriptional control.

• Journal of the Korea Academia-Industrial cooperation Society
• /
• v.21 no.5
• /
• pp.216-222
• /
• 2020

Tumor necrosis factor receptor associated factor 4 (TRAF4) is found to be overexpressed in human breast cancer. It plays a role in cancer metastasis, production of reactive oxygen species, and cell polarity at membranes. The characteristics and functions of TRAF4 in pigs have not yet been identified. As the first step of research, the mRNA sequence of TRAF4 in porcine cells has been determined. To obtain the full-length sequence, rapid amplification of cDNA ends (RACE) has been carried out. Upon cloning, 2,030 bp of nucleotides were found to encode 470 amino acids, and 8 and 12 amino acids were different from those of the human and mouse TRAF4, respectively. The coding region of porcine TRAF4 was shown to be 93% and 90% homologous to human and mouse TRAF4, respectively. qPCR was conducted to determine the relative expression level of TRAF4. TRAF4 expression in pK15 was enhanced by cell-cell contacts. The mRNA levels of CLDN4, OCLN, and TJP1 at 60% and 80% confluency were significantly higher than at 40% confluency. Further, TRAF4 and tight junction-related genes were down-regulated upon treatment with TRAF4 siRNA. Thus, TRAF4 may affect the function of tight junctions in pig.

• The Korean Journal of Pesticide Science
• /
• v.13 no.4
• /
• pp.290-297
• /
• 2009

This research aims to contribute the characterization of acetolactate synthase (Ec 4.1.3.18; ALS) and the resistance mechanism by sequence analysis of ALS gene of the sulfonylurea-resistant and -susceptible Monochoria vaginalis. The ALS gene was obtained from susceptible (S) and resistant (R) M. vaginalis to sulfonylurea herbicides (SUs). The 815 bp the fragment and the genomic DNA sequence coding for acetolactate synthase (ALS) of S and R biotypes of M. vaginalis were cloned and sequenced. Nineteen clones were divided greatly into 4 groups as result of sequencing. The first group was not difference to S type, the second group was amino acid of P197S which found point mutations causing substitution of serine for proline at amino acid 197, the third group was observed greatly other part of 6 places than group 1, and the fourth group appeared the intergrade of group 1 and 3. Therefore, it could be assumed what ALS gene of various types can be one plant. The peptide of the 13 amino acid Domain A region for ALS genes from R biotype of M. vaginalis differed from that of the S biotype by one base substitution at proline codon of Domain A. It could also be confirmed that point mutation of serine for proline at amino acid 197.

• Pediatric Gastroenterology, Hepatology & Nutrition
• /
• v.6 no.2
• /
• pp.112-119
• /
• 2003

Purpose: Helicobacter pylori infection is thought to be correlated with iron-deficiency anemia (IDA) at puberty. The H. pylori feoB gene, a high-affinity ferrous iron transporter, plays a central role in iron acquisition. This study aims to analyze the H. pylori feoB status according to the presence of antral gastritis with or without IDA. Methods: Fourteen H. pylori-positive patients aged from 10~18 years were categorized into subgroups based on the presence or absence of IDA. Eight patients had IDA, and the other six showed normal hematological findings. Genomic DNA was isolated from cultured H. pylori. Five sets of primers were used for PCR amplification of the feoB gene. The feoB region, 1.93 kb, was generated by linking of the PCR products and sequenced. The feoB gene sequences of H. pylori J99 and 26695 were used to compare with the clinical strains. Sequence comparisons of the feoB regions between the IDA (+) and (-) groups were performed. Results: Sequence analysis of the complete coding region of the feoB revealed 16 sites of polymorphism. Among these, 3 polymorphisms-Glu/Thr254Ala, Ile263Val, and Lys511Gln - were indigenous to Korean strains. Although statistically significant differences appear in 4 sites between IDA (+) and (-), the number of specimens are too low to assess the real differences. Conclusion: The 4 polymorphisms in the feoB gene seem to be related with IDA, but it is unclear yet because of small number of study strains. Further studies are required to prove the correlation of IDA and H. pylori infection.

• Proceedings of the Korean Society for Bioinformatics Conference
• /
• 2000.11a
• /
• pp.21-22
• /
• 2000

Expressed sequence tags (EFTs) are the partial segments of cDNA produced from 5 or 3 single-pass sequencing of cDNA clones, error-prone and generated in highly redundant sets. Advancement and expansion of Genomics made biologists to generate huge amount of ESTs from variety of organisms-human, microorganisms as well as plants, and the cumulated number of ESTs is over 5.3 million, As the EST data being accumulate more rapidly, it becomes bigger that the needs of the EST analysis tools for extraction of biological meaning from EST data. Among the several needs of EST analyses, the extraction of protein sequence or functional motifs from ESTs are important for the identification of their function in vivo. To accomplish that purpose the precise and accurate identification of the region where the coding sequences (CDSs) is a crucial problem to solve primarily, and it will be helpful to extract and detect of genuine CD5s and protein motifs from EST collections. Although several public tools are available for EST analysis, there is not any one to accomplish the object. Furthermore, they are not targeted to the plant ESTs but human or microorganism. Thus, to correspond the urgent needs of collaborators deals with plant ESTs and to establish the analysis system to be used as general-purpose public software we constructed the pipelined-EST analysis system by integration of public software components. The software we used are as follows - Phred/Cross-match for the quality control and vector screening, NCBI Blast for the similarity searching, ICATools for the EST clustering, Phrap for EST contig assembly, and BLOCKS/Prosite for protein motif searching. The sample data set used for the construction and verification of this system was 1,386 ESTs from human intrathymic T-cells that verified using UniGene and Nr database of NCBI. The approach for the extraction of CDSs from sample data set was carried out by comparison between sample data and protein sequences/motif database, determining matched protein sequences/motifs that agree with our defined parameters, and extracting the regions that shows similarities. In recent future, in addition to these components, it is supposed to be also integrated into our system and served that the software for the peptide mass spectrometry fingerprint analysis, one of the proteomics fields. This pipelined-EST analysis system will extend our knowledge on the plant ESTs and proteins by identification of unknown-genes.