Establishment and Applications of Small RNA High-throughput Sequencing Analysis Platform

• Main Authors: Huang, Po-Jung, 黃柏榕
• Other Authors: Lyu, Ping-Chiang
• Format: Others
• Language: en_US
• Published: 2011
• Online Access: http://ndltd.ncl.edu.tw/handle/14671378623438405565

博士 === 國立清華大學 === 生物資訊與結構生物研究所 === 99 === MicroRNAs (miRNAs) are a class of extensively studied small RNA population that plays a critical role in eukaryotic gene regulation. In this study, an automated multiple-task web service DSAP was designed to provide a total solution to analyzing deep-sequencing small RNA datasets generated by next-generation sequencing technology. The input data will go through four analysis steps in DSAP: (i) cleanup: removal of adaptors and poly-A/T/C/G/N nucleotides; (ii) clustering: grouping of cleaned sequence tags into unique sequence clusters; (iii) non-coding RNA (ncRNA) matching: sequence homology mapping against a transcribed sequence library from the ncRNA database Rfam (http://rfam.sanger.ac.uk/); and (iv) known miRNA matching: detection of known miRNAs in miRBase (http://www.mirbase.org/) based on sequence homology. The expression levels corresponding to matched ncRNAs and miRNAs are summarized in multi-color clickable bar charts linked to external databases. DSAP is also capable of displaying miRNA expression levels from different jobs using a log2-scaled color matrix. Furthermore, a cross-species comparative function is also provided to show the distribution of identified miRNAs in different species as deposited in miRBase. In order to validate the functions of DSAP, we used deep-sequencing technology to investigate the miRNA expression pattern in four deep-branching unicellular flagellates: Giardia lamblia, Trichomonas vaginalis, Tritrichomonas foetus, and Pentatrichomonas hominis. In addition to the known miRNAs that have been described in G. lamblia and T. vaginalis, we identified 14 ancient animal miRNA families and 13 plant-specific families from the four unicellular flagellates. Bioinformatics analysis also identified four novel miRNA candidates with reliable precursor structures derived from mature tRNAs. Our results indicated that miRNAs are likely to be a general feature for gene regulation throughout unicellular and multicellular eukaryotes and some of them may derive from unconventional ncRNAs such as snoRNA and tRNA.