| Budget Amount *help |
¥6,300,000 (Direct Cost: ¥6,300,000)
Fiscal Year 2000: ¥2,900,000 (Direct Cost: ¥2,900,000)
Fiscal Year 1999: ¥3,400,000 (Direct Cost: ¥3,400,000)
|
|---|
| Research Abstract |
It is estimated that the origin of the motility of the outer hair cell (OHC) in response to electrical stimulation is the conformational change of the motor protein in the lateral wall of the cell. However, this mechanism has not been clarified yet. Therefore, in this study, firstly, the organ of Corti was removed from the guinea pig and the organ of Corti cDNA library was constructed. Then, after each clone was sequenced, an attempt was made to identify the motor protein by analyzing the sequence and tissue expression. As a result, two putative novel genes which might play an important role in the cochlea were found. However, P.Dallos et al. identified the gene that codes for the motor protein and designated it "Prestin" (Nature 405 : 149-155, 2000). Therefore, after that, based on the base sequences of prestin opened to the public web site, an attempt was made to clone gerbil prestin cDNA by polymerase chain reaction (PCR). The results are as follows :
1. As the full length coding sequence of prestin cDNA is quite long and it is difficult to obtain it directly from the gerbil cochlear cDNA, firstly, four fragments of prestin were obtained. Then, these fragments were combined by PCR.As a result, the full length coding sequence of prestin cDNA is obtained.
2. The combining method is useful to clone long cDNA by PCR.In this method, in order to exclude an error in the combined region, it is important to consider the reaction condition of combining method, e.g., the annealing temperature, time, and so on.
|
