国立屏东教育大学97学年度研究所硕士班入学考试.doc

國立屏東教育大學97學年度研究所碩士班入學考試 科學文獻判讀 試題 （應用化學暨生命科學系碩士班 生物組與化學組） ※請注意：1.本試題共三頁。 2.答案須寫在答案卷，否則不予計分。 問答題（共100分） 一、Paragraph I 本文轉載自PNAS (2008) 105: 5809–5814 Gene knockout is the most powerful tool for determining gene function or permanently modifying the phenotypic characteristics of a cell. Existing methods for gene disruption are limited by their efficiency, time to completion, and/or the potential for confounding off-target effects. Here, we demonstrate a rapid single-step approach to targeted gene knockout in mammalian cells, using engineered zinc-finger nucleases (ZFNs). ZFNs can be designed to target a chosen locus with high specificity. Upon transient expression of these nucleases the target gene is first cleaved by the ZFNs and then repaired by a natural—but imperfect—DNA repair process, nonhomologous end joining. This often results in the generation of mutant (null) alleles. As proof of concept for this approach we designed ZFNs to target the dihydrofolate reductase (DHFR) gene in a Chinese hamster ovary (CHO) cell line. We observed biallelic gene disruption at frequencies 1%, thus obviating the need for selection markers. Three new genetically distinct DHFR?/? cell lines were generated. Each new line exhibited growth and functional properties consistent with the specific knockout of the DHFR gene. Importantly, target gene disruption is complete within 2–3 days of transient ZFN delivery, thus enabling the isolation of the resultant DHFR?/? cell lines within 1 month. These data demonstrate further the utility of ZFNs for rapid mammalian cell line engineering and establish a new method for gene knockout with application to reverse genetics, functional genomics, drug discovery, and therapeutic recombinant protein production. Question: 請嘗試說明ZFNs之設計方式與用途。（20％） 請說明內文中以何種實驗証據證明ZFNs之效用。（10％） 請嘗試說明ZFNs未來之應用願景。（10％） 二、Paragraph II Canada intends to ban the import and sale of plastic polycarbonate baby bottles, which would be the first such step in the world, Health Minister