跳躍子flea插入let-7 complex 基因座可能導致mir-100之低表現量，進而造成果蠅存活率降低和發育遲緩

摘要

　　miRNA是小片段的RNA分子，約含21至23個鹼基，可藉由與mRNA的互補配對，抑制蛋白質轉譯。而miRNA cluster，可藉由poly-cistronic轉錄方式使miRNA表現。在let-7 cluster裡包含miR-100、let-7、miR-125這三個miRNA，這個cluster在前人的研究中被證實會在果蠅幼蟲轉變成蛹時表現。但是，一個以microarray的方式偵測miRNA表現的研究中，發現Canton S (CS-UCI)品系中miR-100在前蛹期表現量較其它miRNA或其它品系miR-100表現量較低之現象。本研究以該研究所使用之品系為對象，並以Oregon R (OR-UCI)和得自於長庚大學皮海薇老師實驗室的Canton S (CS-PI)探討miR-100表現量異於同cluster之其他miRNA的原因。經由一系列PCR實驗，發現在CS-UCI的mir-100與let-7區間有一段插入的片段，定序後進行BLAST序列比對，發現這個插入片段有98%的序列與flea跳躍因子相似，因此推測flea可能造成miR-100的低表現量。在不同溫度的蛹存活率測試中，觀察CS-UCI的蛹存活率，於高溫環境下顯著較低，因此推測miR-100可能具有影響蛹於高溫環境中耐受力的功能，這一個獨特的現象，讓我們可以了解miR-100在果蠅生長中扮演的角色。

關鍵詞：miRNA、cluster、mir-100、let-7 complex、果蠅發育

Hypo-expression of mir-100, reduced survival rate, and delayed development in Drosophila melanogaster may result from the insertion of flea transposon in let-7 complex locus

ABSTRACT

　　MicroRNAs (miRNAs) are 21~23nt long RNAs, which regulate gene expression at post-transcriptional level by inhibiting the translation of target mRNAs through complementary pairing. Clustered miRNAs, such as let-7 cluster, are suggested to exhibit coordinate expression via a poly-cistronic transcription or inhibit translation. Let-7 cluster, consisting of miR-100, let-7, and miR-125, are increasingly expressed during larva-to-pupa transition in Drosophila melanogaster. However, the expression level of miR-100 was show to be lower than those of let-7 and miR-125 in Canton S strain as well as those of miR-100 in other strains in previous studies. In this study, we investigated the cause and the effects of low expression level of miR-100 in Canton S strain from UCI (CS-UCI) by using Oregon R (OR) strain and Canton S strains from Dr. Hai-wei Pi’s lab (CS-PI) as control strains. In qRT-PCR experiments, the expression level of miR-100 was confirmed to be low in CS-UCI but high in CS-PI and OR. A series of PCR experiments in genomic DNA revealed the presence of a flea element between mir-100 and let-7 in CS-UCI but not in CS-PI, suggesting the lower expression level of miR-100 might result from this insertion. Base on our preliminary tests, CS-UCI pupae showed significantly lower survival rate at 30℃ than the other two strains, suggesting miR-100 might involve in buffering heat stress during pupal development. The detail molecular mechanism on low expression level of miR-100 and its phenotypic effects in Drosophila development require further investigation.

Key words： microRNAs、miRNA cluster、mir-100、let-7 Complex、Drosophila development

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