An attempt of application Myo D transfected skin fibroblasts of patients to enhance gene expression and to analyze muscle specific Na channel proteins.
| Project/Area Number |
07457129
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
内科学一般
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| Research Institution | OKINAKA MEMORIAL INSTITUTE FOR MEDICAL RESEARCH |
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Principal Investigator |
SHISHIBA Yoshimasa OKINAKA MEMORIAL INSTITUTE, 主任研究員 (00072596)
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| Co-Investigator(Kenkyū-buntansha) |
TAKAHASHI Kunitaro MEIJI COLLEGE OF PHARMACY, 分子生物学, 教授 (10010034)
清水 多恵子 (財)冲中記念成人病研究所, 主任研究員
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| Project Period (FY) |
1995 – 1997
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| Project Status |
Completed (Fiscal Year 1997)
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| Budget Amount *help |
¥4,700,000 (Direct Cost: ¥4,700,000)
Fiscal Year 1997: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1996: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1995: ¥2,300,000 (Direct Cost: ¥2,300,000)
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| Keywords | Hypocalcemia / Hypercalcemia / Ion channel / Myo D gene / Myosin / Fibroblast / Na channel / 皮膚線維芽細胞 |
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| Research Abstract |
Purpose : Our goal of the research is to transform human fibroblasts to myoblasts by transfecting MyoD gene. Such transfected and successfully transformed cells would give us a very handy tool to examine genetic abnormality of muscle-specific praoteins such as ryanodine-receptor protein or TTX-sensitive Na channel. Such tranformed cells would also allow us to examine physiological lproperties of those channel proteins.
Materials and methods : As our rigorous attempts to transfect MyoD gene into human fibfoblasts employing Cap, lipofectine, or lipofectamine, we adopted animal cell-line which has higher rate of transfection efficiency and examined basic requirements to induce TTX-sensitive Na channel employing a sensitive method to detect the channel mRNA.
Results : One of the collaborator, Dr.Okamura succeeded to modify RT-PCR to be sensitive nough to detect changes in TTX-insensitive Na channel expression according to the development of motor neuron in isolated blastomeres of Halocynthia roretzi. Drs.Takahashi and Tanaka have been engaged in developing a new method to detect transfection efficiency directly under the microscope. They arae trying to establish a Myo-DGFP fusion protein and examine the expression in an in-vitro culture system. Although the attempts are currently under the progress, they have already successfully praoduced a fusion protein which connects functional channel protein and also succeeded in expressing the fusion protein in an in-vitro system. By employing these methods, it is feasible to establish a clinically important in vitro system to study transformed skin fibroblasts as a surrogate of muscle biopsy specimen.
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Report
(4 results)
Research Products
(10 results)
