炭疽芽胞杆菌基因芯片探针文库的构建

为制备炭疽芽胞杆菌的基因芯片探针文库，以炭疽芽胞杆菌毒素质粒pX01和荚膜质粒pX02为原材料，用Sau3A I酶切pX01和pX02质粒DNA，Taq DNA聚合酶72℃补平加A，经AT克隆，PCR初步鉴定筛选出炭疽质粒片段的阳性克隆．DNA自动分析仪对克隆片段进行序列测定；用生物信息学方法对其片段进行同源性分析;并将克隆的探针打印于玻片上，制备成炭疽芽胞杆茵基因芯片，与炭疽杆茵质粒DNA样品进行初步芯片杂交的实验，杂交实验的阳性率达到了90％以上，证明大部分克隆探针属于炭疽芽胞杆菌．炭疽芽胞杆菌基因芯片探针文库的构建为基因芯片探针的制备摸索出一条简便、高效、可行的方法．

Construction of Library for Bacillus anthracis Microarray Probes

To prepare the microarray probes of Bacillus anthracis,pX01 and pX02 of B.anthracis were digested with restriction endonuclease Sau3A I.Then the resulting fragments were cloned into the pMD18 T Vectors.To identify the positive clones,a pair of specific vector primers was used to amplify the inserted fragments.The DNA sequences were determined with 377 DNA Sequencer,and the gene sequences were analyzed with bioinformatical software.A DNA microarray of B.anthracis was prepared by printing the probes cloned on a superamine modified glass slide and also was hybridized with B.anthracis plasmids.The sequence analysis and preliminary hybridization results showed that over 90%fragments cloned were specific gene fragments of B.anthracis plasmids,and also some chromosome fragments were included.It is an effective,quick and simple way to prepare microarray probes by way of restriction endonuclease digestion and AT clone.