Childs J D, Birnboim H C
J Virol. 1975 Sep;16(3):652-61. doi: 10.1128/JVI.16.3.652-661.1975.
A method for the electrophoresis of intact bacteriophage T4D particles through polyacrylamide gels has been developed. It was found that phage particles will migrate through dilute polyacrylamide gels (less than 2.1%) in the presence of a low concentration of MgCl2. As few as 5 x 10(9) phage particles can be seen directly as a light-scattering band during the course of electrophoresis. The band can also be detected by scanning gels at 260 to 265 nm or by eluting viable phage particles from gel slices. A new mutant (eph1) has been identified on the basis of its decreased electrophoretic mobility compared with that of the wild type; mutant particles migrated 14% slower than the wild type particles at pH 8.3 and 35% slower at pH 5.0. The isoelectric points of both the wild type and eph1 mutant were found to be between pH 4.0 and 5.0. Particles of T4 with different head lengths were also studied. Petite particles (heads 20% shorter than normal) migrated at the same rate as normal-size particles. Giant particles, heterogenous with respect to head length (two to nine times normal), migrated faster than normal-size particles as a diffuse band. This diffuseness was due to separation within the band of particles having mobilities ranging from 8 to 35% faster than those of normal-size particles. These observations extend the useful range of polyacrylamide gel electrophoresis to include much larger particles than have previously been studied, including most viruses.
已开发出一种使完整的噬菌体T4D颗粒在聚丙烯酰胺凝胶中进行电泳的方法。研究发现,在低浓度MgCl₂存在的情况下,噬菌体颗粒能够在稀释的聚丙烯酰胺凝胶(小于2.1%)中迁移。在电泳过程中,低至5×10⁹个噬菌体颗粒可直接作为光散射带被观察到。该光散射带也可通过在260至265nm波长下扫描凝胶或从凝胶切片中洗脱活的噬菌体颗粒来检测。基于其与野生型相比电泳迁移率降低,鉴定出一种新的突变体(eph1);在pH 8.3时,突变体颗粒的迁移速度比野生型颗粒慢14%,在pH 5.0时慢35%。野生型和eph1突变体的等电点均在pH 4.0至5.0之间。还研究了具有不同头部长度的T4颗粒。小型颗粒(头部比正常颗粒短20%)的迁移速度与正常大小颗粒相同。头部长度不均一(为正常长度的2至9倍)的巨型颗粒作为一条弥散带,其迁移速度比正常大小颗粒快。这种弥散是由于带内迁移率比正常大小颗粒快8%至35%的颗粒发生了分离。这些观察结果扩展了聚丙烯酰胺凝胶电泳的适用范围,使其能够用于研究比以往研究的颗粒大得多的颗粒,包括大多数病毒。