• Journal of Microbiology
• /
• 제42권4호
• /
• pp.361-364
• /
• 2004

Internal ribosome entry site (IRES) of the hepatitis C virus (HCV) is known to be essential for HCV replication and most conserved among HCV variants. Hence, IRES RNA is a good therapeutic target for RNA-based inhibitors, such as ribozymes. We previously proposed a new anti-HCV modulation strategy based on trans-splicing ribozymes, which can selectively replace HCV transcripts with a new RNA that exerts anti-HCV activity. To explore this procedure, sites which are accessible to ribozymes in HCV IRES were previously determined by employing an RNA mapping method in vitro. In this study, we evaluate the intracellular accessibility of the ribozymes by comparing the trans-splicing activ­ities in cells of several ribozymes targeting different sites of the HCV IRES RNA. We assessed the intra­cellular activities of the ribozymes by monitoring their target-specific induction degree of both reporter gene activity and cytotoxin expression. The ribozyme capable of targeting the most accessible site iden­tified by the mapping studies then harbored the most active trans-splicing activity in cells. These results suggest that the target sites predicted to be accessible are truly the most accessible in the cells, and thus, could be applied to the development of various RNA-based anti-HCV therapies.

• 콘크리트학회논문집
• /
• 제12권6호
• /
• pp.43-50
• /
• 2000

In this paper, a new splicing method was applied to design the girder section of bridges with the span length of 25m, 30m, 35m, 40m and 45m. A U-type precast prestressed section was also determined for each bridge. Additionally, the sectional area, beam depth and Guyon's efficiency factor of the spliced U-type sections in each span were analyzed in comparison with the present I-type PSC bridges. As a result, in spite of an increase of 31%∼50% in the sectional areas compared with the I-type precast girders, the spliced U-type the beam depth of the spliced U-type girder was designed as 2,050 mm compared with the I-type precast girder of 2,600mm in a 40m span bridge. The sectional efficiency factors of the spliced U-type sections were analyzed as 0.76∼0.99. It shows that the spliced U-type sections ar of a superior structural efficiency in contrast to the average sectional efficiency factor of 0.66 value in the I-type girders.

• BMB Reports
• /
• 제40권1호
• /
• pp.15-21
• /
• 2007

Breast cancer is the most common malignancy among women, and mutations in the BRCA1 gene produce increased susceptibility to these malignancies in certain families. In this study, the forward 1-13 exons of breast cancer associated gene BRCA1 were cloned from breast cancer cell line ZR-75-30 by RT-PCR method. Sequence analysis showed that nine BRCA1 splice forms were isolated and characterized, compared with wild-type BRCA1 gene, five splice forms of which were novel. These splice isoforms were produced from the molecular mechanism of 5' and 3' alternative splicing. All these splice forms deleting exon 11b and the locations of alternative splicing were focused on two parts:one was exons 2 and 3, and the other was exons 9 and 10. These splice forms accorded with GT-AG rule. Most these BRCA1 splice variants still kept the original reading frame. Western blot analysis indicated that some BRCA1 splice variants were expressed in ZR-75-30 cell line at the protein level. In addition, we confirmed the presence of these new transcripts of BRCA1 gene in MDA-MB-435S, K562, Hela, HLA, HIC, H9, Jurkat and human fetus samples by RT-PCR analysis. These results suggested that breast cancer associated gene BRCA1 may have unexpectedly a large number of splice variants. We hypothesized that alternative splicing of BRCA1 possibly plays a major role in the tumorigenesis of breast and/or ovarian cancer. Thus, the identification of cancer-specific splice forms will provide a novel source for the discovery of diagnostic or prognostic biomarkers and tumor antigens suitable as targets for therapeutic intervention.

• 한국전산구조공학회논문집
• /
• 제15권2호
• /
• pp.251-259
• /
• 2002

부재의 일정구간에 설치된 긴장재의 긴장작업과 이미 긴장되어 있는 상태의 긴장재를 이완시키므로써 의도적으로 유도되는 2차 모멘트를 이용한 새로운 연속화 공법이 개발되었다. 본 공법에서는 이들 긴장재의 긴장과 이완공정이 가장 핵심이 되는 작업이 되며, 실용적인 측면에서는 그 크기와 순서를 결정하는 것이 주된 과제가 될 것이다. 본 논문에서는 이 과정에서 필요한 최적의 긴장과 이완 비율의 조절 과정에 관하여 연구하였다. 그 결과, 긴장작업과 이완작업은 먼저 긴장작업부터 시작하여 3단계에 걸쳐 점진적으로 실시하는 것이 구조의 안전과 시공성을 위해 타당한 방법으로 나타났다. 그리고, 민감도 분석을 통하여 결정되는 선행 긴장작업의 긴장력 크기는 최소와 최대의 비율범위를 동시에 만족하는 값으로 결정되어야 하나, 시공의 효율성을 위해서 필요할 패는 최소 긴장비율만을 하한치로 하여 결정하여도 큰 문제는 없다.

• Advances in concrete construction
• /
• 제6권5호
• /
• pp.509-525
• /
• 2018

In this paper the application of forging process as benefit technique in Reinforced Concrete (RC) beam bars and comparison to lap splices was experimentally investigated with four concrete beam specimens with same dimensions and reinforcement details. The reference specimen was with no splices and the other three beams were with different splices (100% forging in the middle, 50% forging, and 100% lap splices in the middle). Beams were tested with the four points load system. Experimental test results indicated that using forging process as new bar splicing method can have high effects on increasing ductility and energy dissipation of concrete structures. It also proved that this method increased the flexural rigidity, energy absorption, and ductility of the RC beams. And also this research results showed that the flexural capacity and ductility of the beam with 50% forging were respectively increased up to 10% and 75% comparing to that of reference specimen, but the energy absorption of this beams was decreased up to 27%. The ductility of beam with 50% forging was increased up to 25% comparing the ductility of beam with 100% forging.

• 한국콘크리트학회:학술대회논문집
• /
• 한국콘크리트학회 1997년도 가을 학술발표회 논문집
• /
• pp.797-802
• /
• 1997

The current study is a part of series o research about the development of new superstructure system to overcome the engineering problems in the design of bridges of 30m to 45m in span length using the existing bridge systems. The basic concept of new system is the continuation of adjacent tow simple spans composed of the precast prestressed concrete U-type sections. The partial post tensioning method is applied to create the continuity. In this study, the new technique was introduced and applied with an example design of tow span of 40m in span length to find the possibility for practical application as the feasibility study. The obtained results show that the new splicing method is expected to offer significant economical and serviceability advantages.

• Journal of Microbiology and Biotechnology
• /
• 제10권5호
• /
• pp.690-693
• /
• 2000

Efficient intron deletion with the correct splicing of the two exons of the human proinsulin gene was accomplished by a novel stepwise method using genomic DNA [5]. The two exons were separately amplified in two steps, using the second step primers that incorporated additional bases complementary to the other exon. The fragments were combined in a third PCR reaction. Cloning and sequencing of the PCR product demonstrated the correct splicing of the two exons. Expression studies, using the pET9a vector, revealed a protein band with the correct size with respect to human proinsulin as confirmed by SDS-PAGe and Western blot. Proinsulin concentration was estimated to be around 200 mg per liter culture, expressed as inclusion bodies. Protein secretion to the culture medium and periplasmic space was achieved by cloning in the pEZZ18 vector.

• Journal of Information Processing Systems
• /
• 제16권1호
• /
• pp.120-131
• /
• 2020

In order to solve the problem of point clouds coordinate conversion of non-directional scanners, this paper proposes a basic Rodrigues rotation method. Specifically, we convert the 6 degree-of-freedom (6-DOF) rotation and translation matrix into the uniaxial rotation matrix, and establish the equation of objective vector conversion based on the basic Rodrigues rotation scheme. We demonstrate the applicability of the new method by using a bar-shaped emboss point clouds as experimental input, the three-axis error and three-term error as validate indicators. The results suggest that the new method does not need linearization and is suitable for optional rotation angle. Meanwhile, the new method achieves the seamless splicing of point clouds. Furthermore, the coordinate conversion scheme proposed in this paper performs superiority by comparing with the iterative closest point (ICP) conversion method. Therefore, the basic Rodrigues rotation method is not only regarded as a suitable tool to achieve the conversion of point clouds, but also provides certain reference and guidance for similar projects.

• 한국가축번식학회지
• /
• 제18권3호
• /
• pp.199-206
• /
• 1994

본 연구는 생쥐 배 발생과정의 상이한 발현을 RT-PCR법에 의해 무작위 증폭함으로서 새로운 유전자를 손쉽게 크로닝하기 위해 수행되었다. mRNA의 상이한 display법은 Ling 과 Pardee (Science 257, 1992)에 의해 개발되었으며, 최근 Zimermann과 Schultz (PNAS USA 91, 1994)에 의해 재증명되었다. 이 방법은 특정 유전자의 일시적 발현의 변화가 maternal 제어로부터 접합체 제어로의 이행에 따른 발현전이, 다정자 침입과 단일 정자 침입에 의한 배발생의 기능적 차이, 성공적으로 부화한 배반포기 배와 부화에 실패한 배반포기 배에서의 발현의 차이는 물론 세포주기에 따른 유전자 발현 양식의 변화에 따른 새로운 유전자의 크로닝을 가능케 한다. 이 방법에 의해, 2세포기 특이 발현 유전자를 크로닝 하였으며, 이 유전자는 EcoRI제한 효소 처리후 Southern blot을 행한 결과 약 15kb genomic size를 가진 것으로 나타났다. 이 새로운 유전자는 간장 특이적 발현을 나타내었다. 또한, 적어도 2개의 mRNA가 존재하였으며, 이는 RNA splicing에 의한 것으로 추정되었다. (PCR, RT-PCR, cloning, preimplantation, mouse)

• 원예과학기술지
• /
• 제35권3호
• /
• pp.323-332
• /
• 2017

Homology-specific transcriptional and post-transcriptional silencing, an intrinsic mechanism of gene regulation in most eukaryotes, can be induced by anti-sense, co-suppression, or hairpin-based double-stranded RNA. Hairpin-based RNA interference (RNAi) has been applied to analyze gene function and genetically modify crops. However, RNAi vector construction usually requires high-cost cloning steps and large amounts of time, or involves methods that are protected by intellectual property rights. We describe a more effective method for generating intron-spliced RNAi constructs. To produce intron-spliced hairpin RNA, an RNAi cassette was ligated with the first intron and splicing sequences of the Brassica rapa ssp. pekinensis histone deacetylase 1 gene. This method requires a single ligation of the PCR-amplified target gene to SpeI-NcoI and SacI-BglII enzyme sites to create a gene-specific silencing construct. We named the resulting binary vector system pKHi and verified its functionality by constructing a vector to silence DIHYDROFLAVONOL 4-REDUCTASE (DFR), transforming it into tobacco plants, and confirming DFR gene-silencing via PCR, RT-qPCR, and analysis of the accumulation of small interfering RNAs. Reduction of anthocyanin biosynthesis was also confirmed by analyzing flower color of the transgenic tobacco plants. This study demonstrates that small interfering RNAs generated through the pKHi vector system can efficiently silence target genes and could be used in developing genetically modified crops.