| Budget Amount *help |
¥5,070,000 (Direct Cost: ¥3,900,000、Indirect Cost: ¥1,170,000)
Fiscal Year 2016: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Fiscal Year 2015: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
Fiscal Year 2014: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
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| Outline of Final Research Achievements |
In our previous studies, we established a method to analyze cells collected by fluorescence-activated cell sorting (FACS), named mRNA quantification after FACS (FACS-mQ). In FACS-mQ, cells are labeled with a fluorescence dyes, and then cells sorted by FACS are examined by analyzing their gene expression profile. Using the previous FACS-mQ protocol, it was not possible to apply some methods, such as amplification of RNAs using T7 promoter, due to RNA degradation. In this study, we tried to modify the FACS-mQ protocol to prevent RNA degradation during FACS. Addition of RNase inhibitor and DTT to some buffers used in FACS-mQ resulted in a high RNA integrity number after FACS. Furthermore, no significant differences in the images of flow cytometry were observed between specimens with or without RNase inhibitor and DTT. Thus, this modification to the FACS-mQ protocol assured the quality of RNAs in cells recovered by FACS.
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