Mathew M K, Smith C L, Cantor C R
Department of Genetics and Development, College of Physicians and Surgeons, Columbia University, New York, New York 10032.
Biochemistry. 1988 Dec 27;27(26):9204-10. doi: 10.1021/bi00426a019.
Pulsed-field gel electrophoresis (PGF) subjects DNA alternately to two electrical fields to resolve DNA ranging from 10,000 base pairs (10 kb) to 10,000 kb in size. The separations are quite sensitive to a variety of experimental variables. This makes it critical to have a wide range of reliable size standards. A technique is described for preparing mixtures of bacteriophage DNA oligomers that span a size range from monomer to more than 30-mer. The relationship between size and mobility of oligomers of different bacteriophage DNA monomers is generally self-consistent. Thus, these samples can serve as primary length standards for DNAs ranging from 10 kb to more than 1500 kb. They have been used to estimate the size of the chromosomal DNAs from various Saccharomyces cerevisiae strains and to test the effect of gel concentration and temperature on PFG. DNA resolution during PFG is slightly improved in agarose gels with small pore sizes, in contrast to continuous electrophoresis where the opposite is observed. PFG mobility is surprisingly sensitive to changes in the running temperature.
脉冲场凝胶电泳（PGF）使DNA交替暴露于两个电场中，以分离大小从10,000个碱基对（10 kb）到10,000 kb的DNA。这种分离对多种实验变量非常敏感。这使得拥有广泛的可靠大小标准至关重要。本文描述了一种制备噬菌体DNA寡聚体混合物的技术，该混合物的大小范围从单体到超过30聚体。不同噬菌体DNA单体的寡聚体大小与迁移率之间的关系通常是自洽的。因此，这些样品可以作为10 kb至超过1500 kb的DNA的主要长度标准。它们已被用于估计各种酿酒酵母菌株染色体DNA的大小，并测试凝胶浓度和温度对脉冲场凝胶电泳的影响。与连续电泳相反，在小孔径琼脂糖凝胶中脉冲场凝胶电泳期间的DNA分辨率略有提高，在连续电泳中观察到的情况则相反。脉冲场凝胶电泳迁移率对运行温度的变化出奇地敏感。
