• Genomics & Informatics
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• v.2 no.1
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• pp.45-52
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• 2004

The self-splicing group I intron from Tetrahymena thermophila has been demonstrated to perform splicing reaction with its substrate RNA in the trans configuration. In this study, we explored the potential use of the trans-splicing group I ribozymes to replace a specific RNA with a new RNA that exerts any new function we want to introduce. We have chosen thymidine phosphorylase (TP) RNA as a target RNA that is known as a valid cancer prognostic factor. Cancer-specific expression of TP RNA was first evaluated with RT-PCR analysis of RNA from patients with gastric cancer. We determined next which regions of the TP RNA are accessible to ribozymes by employing an RNA mapping strategy, and found that the leader sequences upstream of the AUG start codon appeared to be particularly accessible. A specific ribozyme recognizing the most accessible sequence in the TP RNA with firefly luciferase transcript as a 3' exon was then developed. The specific trans-splicing ribozyme transferred an intended 3' exon tag sequence onto the targeted TP transcripts, resulting in a more than two fold induction of the reporter activity in the presence of TP RNA in mammalian cells, compared to the absence of the target RNA. These results suggest that the Tetrahymena ribozyme can be a potent anti-cancer agent to modify TP RNAs in tumors with a new RNA harboring anti-cancer activity.

• The Plant Pathology Journal
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• v.18 no.4
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• pp.187-191
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• 2002

This paper describes the cloning and sequence analysis of the 5'-terminal region and full-length cDNA production of genomic RNA of Lily symptomless virus (LSV), a Species Of the genus Carlavirus. A sing1e DNA band about 600 bp harboring the 5'-end of genomic RNA of the virus was successfully amplified by reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE), and was cloned for nucleotide sequence determination. Sequence analysis of selected RACE cDNA clones revealed that the LSV 5'non-translated region consists of 67 nucleotides long of AT rich stretch followed GC rich from the 5'-end. To produce full-length cDNA products for the viral genomic RNA, a set of LSV-specific primers could be designed based on the obtained sequence in this study and the known sequences of 3'-terminal region for the virus. Full-length cDNA copies of LSV, an 8.4 kb long, were directly amplified by the long-template RT-PCR technique from the purified viral genomic RNA samples. This full-length cDNA copies were analyzed by restriction mapping. The molecules produced in this study can be useful for the production of in vitro infectious cDNA clone, as well as, for the completion of genomic RNA sequence and genome structure for the virus.

• Korean Journal of Microbiology
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• v.35 no.3
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• pp.211-217
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• 1999

Effects of subsh-ate RNA configuration on the tians-splicing reactcon by group I intron ribozyme of Tetralzynzena thern\ulcornerophila were analyzed with substrate RNAs which have been generated to have very stable structures with stem-loop. RNAinapping strategy was perfo~med in vivo as well as in virro to search the mosl accessible siles to the ~irms-splicing ribozymes in the substrate RNAs. Sequences present in the loop of the target RNAs have shown to be well recognized by and reacted with group I inlron ribozymes while sequences present in the stein do not. Thesc results were confirmed with the experiments of trans-cleavage and rmnssplicing reactmn with ihe specific ribozyines recognizing those sequences. Moreover, sequence analysis of the trans-splicing products have shown that irons-splicing reaction can proceed with high fidelity. In conclusion, the secondary structure of substrate RNAs is one of the most important factors to detemine the ribozyme activity.

• Korean Journal of Medicinal Crop Science
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• v.16 no.3
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• pp.195-198
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• 2008

Karyotypic analysis and FISH (fluorescence in situ hybridization) with 45S and 5S rRNA genes were carried out in Persicaria tinctoria H Gross. The somatic metaphase chromosomes were ranged from 2.25 ${\mu}m$ to 1.50 ${\mu}m$ in length. Chromosome number was 2n = 4x = 40 with the basic number of x = 10. The chromosome complement of the species consisted of 16 pairs of metacentrics (chromososomes 1,2,3,4,6,7,8,9, 10, 11, 12, 13, 15, 18, 19 and 20) and 4 pairs of submetacentrics (chromosome 5, 14, 16 and 17). The karyotype formula was K(2n) = 4x = 32 m + 8 sm. In FISH analysis, three pairs of 45S rRNA gene loci on the terminal region of submetacentrics (chromosomes 5, 16 and 17) and two pairs of 5S rRNA gene loci on the centromeric region of metacentrics (chromosomes 9 and 11) were detected, respectively.

• Proceedings of the Zoological Society Korea Conference
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• 1998.10b
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• pp.344.2-344
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• 1998

No Abstract, See Full Text

• Proceedings of the Korean Society of Applied Pharmacology
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• 1996.11a
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• pp.55-64
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• 1996

mouse chromesome2에 있는 agouti locus는 정상적으로는 털색깔을 조절하는 gene이다. mouse agouti gene은 최근에 cloning 되었고 131 amino acid peptide와 consensus signal peptide를 encode한다고 보고되었다. 이 논문에서 interspecies-DNA hybridization approach를 이용하여 mouse agouti gene의 human homologue를 cloning 하였다. Sequence analysis 결과, 이는 mouse gene에 85% 유사하였고 consensus signal peptide sequence 를 포함하는 132 amino acid를 coding하였다. somatic-cell hybrid mapping pannel과 Fluorescence-in-situ hybridization에 의한 chromosomal mapping을 한 결과, agouti gene은 MODY (maturity onset diabetes of the young), myeloid leukemia locus 등이 위치한 human chromosome 20q 11.2에 mapping 되었다. 성인 tissue로부터 추출한 RNA를 이용한 발현연구에 의하면 human agouti gene은 adipose tissue와 teatis에 발현되었다.

• KSBB Journal
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• v.26 no.2
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• pp.131-138
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• 2011

Some computational approaches are needed for clarifying RNAi sequences, because it takes much time and endeavor that almost of RNAi sequences are verified by experimental data. Incorrectness of RNAi mechanism and other unaware factors in organism system are frequently faced with questions regarding potential use of RNAi as therapeutic applications. Our massive parallelized pair alignment scoring between dsRNA in Genebank and expressed sequence tags (ESTs) in Caenorhabditis elegans Genome Sequencing Projects revealed that this provides a useful tool for the prediction of RNAi induced cosuppression details for practical use. This pair alignment scoring method using high performance computing exhibited some possibility that numerous unwanted gene silencing and cosuppression exist even at high matching scores each other. The classifying the relative higher matching score of them based on GO (Gene Ontology) system could present mapping dsRNA of C. elegans and functional roles in an applied system. Our prediction also exhibited that more than 78% of the predicted co-suppressible genes are located in the ribosomal spot of C. elegans.

• Journal of Microbiology and Biotechnology
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• v.4 no.3
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• pp.183-190
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• 1994

Cell-free extracts prepared from cultured insect cells, Spodoptera. frugiperda, were analyzed for activation of early gene transcription of an insect baculovirus, Autographa californica nuclear polyhedrosis virus (AcNPV). The template DNA used for in vitro transcription assays contained promoter sites for the baculovirus genes that have been classified as immediate early ($\alpha$) or early genes. These genes are located in the HindIII-K/Q region of the AcNPV genome. Nuclei isolated from the AcNPV-infected Spodoptera frugiperda cells were also used for in vitro transcription analysis by RNase-mapping the labeled RNA synthesized from in vitro run-on reaction in the isolated nuclei. The genes studied by this technique were p26 and pl0 genes which were classified as delayed early and late gene, respectively. We found that transcription of the genes from the HindIII-K region was accurately initiated and unique in the whole cell extract obtained from uninfected cells, although abundance of the in vitro transcripts was reverse to that of in vivo RNA. With isolated nuclei transcription of the p26 gene was inhibited by $\alpha$-amanitin suggesting that the p26 gene was transcribed by host RNA polymerase II. However, transcription of the pl0 gene in isolated nuclei was not inhibited by $\alpha$-amanitin, but rather stimulated by the inhibitor. We also found that the synthesis of $\alpha$-amanitin-resistant RNA polymerase was begun before 6 hr p.i., the time point at which the onset of viral DNA replication as well as the appearance of a-amanitin-resistant viral transcripts were detected. These studies give us strong evidence to support the previous data that early genes of AcNPV were transcribed by host RNA polymerease III, while transcription of late genes was mediated at least by a novel $\alpha$-amanitin-resistant RNA polymerase.

• Journal of Sericultural and Entomological Science
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• v.42 no.1
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• pp.14-23
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• 2000

The mitochondrial genome of domesticated silkworm (Bombyx mori) was mapped with five restriction endonucleases (BamHI, EcoRI, HindIII, PstI and XbaI), the entire genome was cloned with HindIII and EcoRI. From the end sequencing results of 5$^1$and 3$^1$region for full genome set of eleven mitochondrial clones, the seven mitochondrial genes (NADH dehydrogenase 6, ATPase 6, ATPase 8, tRN $A^{Lys}$, tRN $A^{Asp}$, tRN $A^{Thr}$ and tRN $A^{Phe}$ of mori were identified on the basis of their nucleotide sequence homology. The nucleotide composition of NADH dehydrogenase 6 was heavily biased towards adenine and thymine, which accounted for 87.76%. On basis of the sequence similarity with published tRNA genes from six insect species, the tRN $A^{Lys}$, tRN $A^{Asp}$ and tRN $A^{Thr}$ were showed stable canonical clover-leaf tRNA structures with acceptible anticodons. However, both the DHU and T$\psi$C arms of tRN $A^{Phe}$ could not form any stable stem-loop structure. The two overlapping gene pairs (tRN $A^{Lys}$ -tRN $A^{ASP}$ and ATPase8-ATPase6) were found from our sequencing results. The genes are encoded on the same strad. ATPase8 and ATPase6 overlaps (ATGATAA) which are a single example of overlapping events between abutted protein-coding genes are common, and there is evidence that the two proteins are transcribed from a single bicistronic message by initiation at 5$^1$terminal start site for ATPase8 and at an internal start site for ATPase6. Ultimately, this result will provide assistance in designing oligo-nucleotides for PCR amplification, and sequencing the specific mitochondrial genes for phylogenetics of geographic races, genetically improved silkworm strains and wild silkworm (mandarina) which is estimated as ancestal of domesticated silkworm.sticated silkworm.

• Journal of Microbiology and Biotechnology
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• v.1 no.4
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• pp.256-260
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• 1991

In B. subtilis, $\alpha$-amylase synthesis is regulated by amyR located directly on the upstream of amyE. Three different amyR alleles have been reported, amyR1, amyR2 and amyR3. Strains bearing the gra-10 mutation which confers derepression for catabolite repression has GlongrightarrowA transition mutation at ＋5 of amyR1. S1 nuclease mapping demonstrated that transcription initiated at 8 bases downstream from the -10 region of putative E$\sigma^{A}$ promoter P1 in amyR1 and gra-10. In amyR2, the major transcription initiatd at the same place and the minor, 10 bases downstream from -10 of P2. The transcript from P2 contributed approximately 15-20% of total amyE mRNA. S1 nuclease protection experiment indicated that amyE mRNA levels corresponded to the rate of synthesis assumed by specific activities of $\alpha$-amylase in culture supernatants, suggesting that $\alpha$-amylase synthesis is regulated at the level of transcription.n.