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    • 2. 发明公开
    • METHOD OF CONSTRUCTING MODIFIED PROTEIN
    • 构建修饰蛋白的方法
    • EP1760149A1
    • 2007-03-07
    • EP05737205.4
    • 2005-05-06
    • PRECISION SYSTEM SCIENCE CO., LTD.
    • TAJIMA, Hideji, PRECISION SYSTEM SCIENCE CO., LTDTAKAHASHI, Masaaki PRECISION SYSTEM SCIENCE CO LTDMIYASHITA, Yukiko, PRECISION SYSTEM SCIENCE CO LTD
    • C12N15/00C12N9/00C12P21/02G01N33/15G01N33/50C07K1/00
    • C12N15/1034C12N15/10C12N15/102C12P21/02
    • DNA derived from a microorganism is prepared from a sample containing the microorganism, degenerate primers are prepared based on an amino acid sequence conserved among known proteins nos. 1 through N all having the same type of function, PCR is performed using the degenerate primers in the presence of the DNA derived from the microorganism, the nucleotide sequence of DNA fragment amplified by the PCR is determined and the amino acid sequence coded for by the DNA fragment is also determined, homology between the amino acid sequence coded for by the DNA fragment and amino acid sequences of known proteins is evaluated, a known protein is discovered which has an amino acid sequence having homology with the amino acid sequence coded for by the DNA fragment, the region of the DNA coding for the discovered known protein which codes for the amino acid sequence having the homology is replaced by the DNA fragment to produce a modified DNA, and the modified DNA is expressed to produce a modified protein, thereby achieving high-throughput and efficient functional analysis and screening of a modified protein.
    • 从含有微生物的样品制备来自微生物的DNA,根据已知蛋白质中保守的氨基酸序列制备简并引物。 1至N都具有相同类型的功能,在来自微生物的DNA的存在下使用简并引物进行PCR,测定通过PCR扩增的DNA片段的核苷酸序列,并确定由该编码的氨基酸序列 也测定DNA片段,评估由该DNA片段编码的氨基酸序列与已知蛋白质的氨基酸序列之间的同源性,发现一种已知的蛋白质,其具有与由该氨基酸序列编码的氨基酸序列具有同源性的氨基酸序列 DNA片段中,编码发现的已知蛋白质的编码具有同源性的氨基酸序列的DNA的区域被DNA片段置换以产生修饰的DNA,并且修饰的DNA被表达以产生修饰的蛋白质,从而实现 高通量和高效的功能分析和筛选改性蛋白。