Ely B, Gerardot C J
Department of Biology, University of South Carolina, Columbia 29208.
Gene. 1988 Sep 7;68(2):323-33. doi: 10.1016/0378-1119(88)90035-2.
The restriction enzyme DraI cleaves the Caulobacter crescentus genome into at least 35 fragments which have been resolved in agarose gels using pulsed-field-gradient gel electrophoresis (PFGE). When digests were performed using DNA from strains containing Tn5 insertion mutations, altered band migrations were observed. Using PFGE with the appropriate pulse times, size differences as small as 2% could be resolved in large fragments. Using this approach, we have constructed a partial physical map of the genome which correlates well with the C. crescentus genetic map and have shown the size of the genome to be approx. 3800 kb. Using hybridization with cloned genes, we have determined the map locations of five previously unmapped genes. In addition, we have shown that PFGE can be used to rapidly determine the map locations of new insertion mutations or the sizes of deletion mutations.
限制性内切酶DraI可将新月柄杆菌基因组切割成至少35个片段，这些片段已通过脉冲场梯度凝胶电泳（PFGE）在琼脂糖凝胶中得到分离。当使用含有Tn5插入突变的菌株的DNA进行酶切时，观察到条带迁移发生了改变。使用具有适当脉冲时间的PFGE，在大片段中可以分辨出小至2%的大小差异。采用这种方法，我们构建了与新月柄杆菌遗传图谱相关性良好的基因组部分物理图谱，并表明基因组大小约为3800 kb。通过与克隆基因杂交，我们确定了五个先前未定位基因的图谱位置。此外，我们还表明PFGE可用于快速确定新插入突变的图谱位置或缺失突变的大小。
