Q热立克次体基因文库的构建

Construction of A Coxiella burnetii Gene Library

Ｑ热立克次体七医株用６种限制性平端内切酶分酶切，电泳回收０．２－７ｋｂ大小ＤＮＡ片段，ＥｃｏＲＩ位点甲基化，加上ＥｃｏＩＲ接头，与去磷酸化λｔｇｌｌＤＮＡ－ＥｃｏＲＩ臂连接，经体外包装后感染Ｅ．ｃｏｌｉＹ１０９０，建成含２．６×１０＾６重组子的表达型基因文加，随机选择的噬菌体ＤＮＡ经ＥｃｏＲＩ酶切鉴定，重组子含大小不等的插入ＤＮＡ片段。

Coxiella burnetii QIYI DNA was partially digested with six kinds of restriction enzyme to yield blunt ends. The digested DNA were electrophoresed on a agarose gel and fragments of approximately 0. 2 to 7 kilobases (Kb) were collected. EcoRI sites of DNA were methylated with EcoRI methylase. Linkers were ligated with DNA. Linkered DNA was passed over a Sephacryl S-400 column to remove unligated linkers and then ligated into dephosphorylated EcoRI-digest-ed λgtll DNA. The DNA was packed into bacteriophage particles and used to infect E. coli Y1090. 2. 17×106 clones which contained 2.16×106 white plaques were obtained.