在實驗室裡將模式生物的特定基因進行突變有助於對該基因的功能做進一步的分析，其中秀麗隱桿線蟲因為其生命週期短、容易培養生長、並且也可作為基因轉殖動物而廣泛應用於功能性遺傳學的研究。數種常用來對線蟲進行基因體編輯的方法包括：化學藥劑、輻射、跳躍轉座子等都可使其基因產生隨機性的突變。較為新穎的基因編輯工具-CRISPR系統可針對指定的目標基因造成DNA雙股螺旋結構斷裂，目前也已經用來產生線蟲的突變株。而要使CRISPR系統能作用於線蟲上通常是以單隻注射的顯微注射法的方式來達成，同樣用來產生線蟲的基因轉殖動物的方法還有微粒子轟擊法。雖然微粒子轟擊法可將質體同時送入多隻線蟲的性腺中，但此方法須使用特定基因突變的蟲株以利於之後的篩選，同時也因此大幅度的延長了整個流程時間。近年來線蟲專用的抗生素篩選系統已有所發展，如果將其與微粒子轟擊法結合，將會大幅縮短篩選的時間。在本研究中我們以CRISPR系統使目標基因的外顯子序列產生突變，成功地產生了unc-119與iglr-2的突變蟲株。為了研究線蟲是否具有模式辨別受器，我們給予iglr-2突變蟲株餵食具有致病性的大腸桿菌，以野生型線蟲作為對照組；實驗結果顯示兩者之間並無顯著差異，推測其原因是此iglr-2突變蟲株是屬於框內缺失突變，對其蛋白質功能表現不具影響；同時我們也成功地結合微粒子轟擊法和抗生素篩選系統，產生出了iglr-2過表現蟲株，同樣給予餵食致病性大腸桿菌，並發現該蟲株比野生型更具有抵抗力。在本研究中我們建立了兩個嶄新的技術平台，希望之後對於iglr-2甚至是其他未知基因的功能性分析能夠有所助益。

Mutations allow gene function to be discovered in lab. Caenorhabditis elegans is highly extensive to functional genetics because of its short lifespan, ease of growth, and can be transformed to generate transgenic animals. Several methods for genome editing have been developed for C. elegans. These include random mutation through chemical reagents, and random insertion of transposon in the whole genome. The development of CRISPR-Cas9 system to create targeted DNA double stranded breaks (DSBs) is now able to provide a simple and efficient tool with which the genome of C. elegans is addressable for precisely engineering. The CRISPR-Cas9 system working in C. elegans expresses from the plasmids via microinjection. Another method to generate transgenic worms is microparticle bombardment. However, the method is limited for specific genetic background worms and may need much time for screening. The antibiotic selection systems developed in recent years speeds up the selection process after bombardment. Here, the unc-119 and iglr-2 mutant worm generated via the CRISPR-Cas9 systems in which the mutation site is in the target exon sequence. For study of the pattern-recognition receptor in C. elegans, we feed the iglr-2 mutants pathogenic E. coli. Nevertheless, there are no difference between the mutant and wild type worms in terms of their survival. The possible interpretation is that the mutation is an one amino acid in-frame deletion and the function of the iglr-2 has not been altered. Also, the iglr-2 overexpression worms generated from combination of the bombardment and antibiotic selection methods exhibit significant resistance to the pathogen. On the technological platforms built in this research, it will be effortless to analyze iglr-2 and other unknown gene.

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