| Summary: | 碩士 === 國立成功大學 === 化學工程學系碩博士班 === 94 === The expression of blue fluorescent protein gene (bfp) and mutated gene (bfp-D7) from bfp by random mutagenesis isolated from Vibrio vulnificus CKM-1 in the giant protoplasts of Escherichia coli XL1B and V. vulnificus CKM-1 were investigated, respectively. In addition, we built a system for quantifying the fluorescent intensity of BFP and screened a high efficiency promoter.
We defined two kinds of fluorescence density, one was specific blue density that was de-unit data from Image-Pro Plus’ color depth ; another’s overall blue density that was from specific blue density multiplied optical density. The overall blue density was able to represent specific blue density normalization and evaluated transcription efficiency in promoter. About promoter efficiency evaluated, we had three kinds of variables including two parameters with eight variations, one was bateria including E.coli XL1-Blue and V. vulnificus CKM-1, one was fluorescence protein including BFP and BFP-D7, another was temperature including 30°C and 25°C. According the diagram of overall blue density varied with time, we found the slope of regression line fitting the curve which range was from the highest fluorescence value to the lowest value. The slope represented promoter efficiency. The results showed that the most adaptable system to detect promoter intensity was E.coli XL1-Blue_pUD7 in 30°C.
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