Transcriptome analysis for tunic cells of the ascidian, Ciona intestinalis

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This study intend to build a molecular biological basis of tunic cells in an ascidian, Ciona intestinalis, which is now an important model organism of EvoDevo research. Although tunic is the main characteristic-organ of this animal, the group of cells which are inlaying of the organ is not really investigated to date.The original plan of this study was made up of two parts of strategy that are (1) decoding of RNAs expressed in the tunic cells and, (2) molecular and cellular study for the tunic cells. The RNA extraction from tunic cells for NGS sequencing were difficult than I have been expected.To avoid this problem, we corrected the original plan in the term as followings; (a) we collected the frozen sample of tunic-organ, (b) we provided the continued requirement-study to isolate the RNAs from tunic cells and (c) we designed a custom microarray for future analysis of gene expression in tunic cells. (See Matsumae et al. 2013 in Section 13.)In order to forward the second of abovestrategies , we were concerned about the cellulose filament which is the main component of tunic as an obstruction for analysis of tunic cells. On the advices of Prof. Hirose who is one of cooperative researcher of this study, we overcame thissecond difficulty. We established a method to isolate the tunic cells from the tunic-organ by means of incubation of tunic with a glass slide. This method allowed us to observe a clear view of the tunic-cells and its nuclei by DAPI-staining. We have verified that two types of tunic-cells are isolated from the crude tunic that contains at least three types of cells by our method. We expect that this method has made possible for variety of analytical approaches including of in situ hybridization and cell culture of C. intestinalis.As for future perspectives, because we have collected the tunic-organ sample for RNA-extraction, I'm planning for decoding the RNA sequences of tunic cells within around next two years.