金屬硫蛋白在大腸桿菌的表達與金屬累積能力測試;Metallothionein expression in Escherichia coli and metal accumulation ability test

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Title:	金屬硫蛋白在大腸桿菌的表達與金屬累積能力測試;Metallothionein expression in Escherichia coli and metal accumulation ability test
Authors:	涂文峰;Tu, Wen-Feng
Contributors:	生命科學系
Keywords:	重金屬;生物復育;金屬硫蛋白;基因工程;heavy metals;biological remediation;metallothionein;genetically engineered bacteria
Date:	2019-07-11
Issue Date:	2019-09-03 14:32:28 (UTC+8)
Publisher:	國立中央大學
Abstract:	在於自然環境中，具有強烈的毒性，易造成人體危害。當生物暴露在重金屬環境時，為處理體內的重金屬會表現金屬硫蛋白，並與重金屬結合降低重金屬的毒性。根據前人文獻，建構並且比較3種形式的持續表現金屬硫蛋白(metallothionein, MT)載體在大腸桿菌中的表現情形，在大腸桿菌中只有單獨表現金屬硫蛋白的情況，因金屬硫蛋白是不穩定的小分子蛋白，因此在大腸桿菌中的表達並不是很穩定；然而當金屬硫蛋白接上綠螢光蛋白(EGFP)或是谷胱甘肽S-轉移酶(GST)時，金屬硫蛋白能在大腸桿菌中穩定表達。而在測試重金屬抗性，確定大腸桿菌能生長於150 ppm Ni2+ 和 Cd2+之培養基。因為大腸桿菌的鎳離子外排系統，使鎳離子不能長時間存在於大腸桿菌內，無法應用在鎳的生物復育，然而在大腸桿菌表達金屬硫蛋白之融合蛋白的情況，在培養24小時後能穩定結合著鎳離子，避免鎳離子在大腸桿菌體內的大量流失，有效吸收鎳離子1.5 mg/g，顯示金屬硫蛋白對於增加鎳的吸收量是有效的。;Heavy metals exist in the natural environment and have strong toxicity, which is easy to cause harm to the human body. When organisms are exposed to heavy metals, metallothioneins combine with heavy metals to reduce the toxicity of heavy metals. The three types of sustained expression of metallothionein (MT) plasmid in Escherichia coli were constructed and compared. In Escherichia coli, only metallothionein was expressed alone, because metallothionein is a small molecule protein not stable, however, metallothionein is attached to green fluorescent protein (EGFP) or glutathione S-transferase (GST), metallothionein can be stably expressed in Escherichia coli. While testing for heavy metal resistance, it was confirmed that Escherichia coli can grow in medium of 150 ppm Ni2+ or Cd2+. Because of the nickel ion efflux system of Escherichia coli, nickel ions cannot be present in Escherichia coli for a long time, and cannot be used for bioremediation of nickel. However, the expression of metallothionein fusion protein in Escherichia coli, after 24 hours, to avoid the massive loss of nickel ions from Escherichia coli, and effectively absorb nickel ions 1.5 mg/g, indicating that metallothionein is effective for increasing the absorption of nickel.
Appears in Collections:	[Graduate Institute of Life Science] Electronic Thesis & Dissertation

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