| Project/Area Number |
13680803
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Developmental biology
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
SHUKUNAMI Chisa Institute for Frontier Medical Sciences. Associate professor, 再生医科学研究所, 助教授 (60303905)
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| Co-Investigator(Kenkyū-buntansha) |
YAMADA Shuich Institute of vVlral Research , Instructor, ウイルス研究所, 助手 (20261133)
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| Project Period (FY) |
2001 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2002: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2001: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Keywords | ChMrP / tenomodulin / chondromodulin-l / dense connective tissue / knockout mice / tendon / anti-angiogenic barrier / angiogenesis |
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| Research Abstract |
Chondromodulin-l related protein (ChM-rP) was cloned as a homologous gene to choondromodulin-l (ChM-l), a cartilage derived angiogenesis inhibitor. We renamed it tenomodulin (TeM) after the name of tenocytes that are longitudinally aligned fibroblastic cells of tendon, one of typical dense connective tissues. Analysis of ChM-l null mice revealed abnormal bone metabolism after birth but no alteration of the onset of primary ossification nor vascular invasion, suggesting an importance for generation of a related gene TeM that is expressed in the overlapping or nieghboring regions positive for ChM-l. TeM genomic clones were isolated from a TT2 embryonic stem cell genomic library. A 1.4 kb genomic fragment was amplified by PCR and subcloned into pPNT at Xba-l and Kpn-l sites. A 5.2 kb genomic fragment containing the flanking region of TeM was ligated to the Xho-l site of the vector. TT2 ES cells were transfected with a targeting vector Inialized with Not l by BioRad Gene Pulser. ES clones were screened by Southern blot analysis or PCR. No positive clone was obtained after screening of 400 ES clones. To improve the transfection efficiency, long arm region was extended to 6.8 kb. Radiation hybrid mapping analysis revealed that TeM is a X-linked gene, suggesting the possibilities of low efficient homologous recombination in ES cells and prevention of germline transmission in chimeras. To avoid this problem, construction of a Cre-loxP targeting vector and knockdown by RNAi are being performed
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