Methods for fabrication of uncultivated genome-probing microarrays by amplifying genome derived from a single microbial cell using multiple displacement amplification

摘要

A method for fabricating genome microarray by amplifying the genome derived from a single cell of uncultivated microorganism by using digital multiple displacement amplification is provided to classify and detect microorganisms accurately without cultivation. A method for fabricating genome microarray comprises the steps of: sampling an uncultivated microorganism from the environment and diluting the sample to divide into single cells; denaturing the cell wall and cell membrane of the single cells of uncultivated microorganism by using lysozyme to obtain the genome; amplifying the genome of denatured uncultivated microorganism through digital multiple displacement amplification(MDA); analyzing the nucleotide sequence of the amplified genome by using one method selected from PCR sequencing, DNA fingerprint analysis, DGGE(denaturing gradient gel electrophoresis) and tRFLP(terminal restriction fragment length polymorphism) and confirming whether the analysis result is indicated by the single chromatogram; and spotting the MDA amplifying product derived from the single cell to a support.