By Kaplan N.P. (ed.), Colowick N.P. (ed.), Wu R. (ed.)
The looks of one other quantity in that very good sequence, tools in Enzymology, is usually a reason for appreciation when you desire to effectively perform a specific method or organize an enzyme or metabolic intermediate with no the tiresome prospect of looking through surprising literature and maybe opting for an unproven process which isn't simply reproduced.
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4 p. H. Seeburg, J. Shine, J. A. Martial, J. D. Baxter, and H. M. Goodman, Nature (London) 270, 486 (1977). 5 A. Efstratiadis, F. C. Kafatos, and T. Maniatis, Cell 10, 279 (1977). 6 R. Breathnach, J. L. Mandel, and P. Chambon, Nature (London) 270, 314 (1977). 7 C. Brack, M. Hirama, R. Lenhard-Schuler, and S. Tonegawa, Cell 15, 1 (1978). 8 H. Temin and S. Mizutani, Nature (London) 226, 1211 (1970). 9 A. Efstratiadis and L. Villa-Komaroff, in "Genetic Engineering" (J. K. Setlow and A. ), Vol. 1, pp.
If the tailed vector is not affinity purified on oligo(dA)-cellulose, the background in the transformations is unacceptably high (50% or greater), probably due to the presence of undigested or untailed plasmid. One pass lowers the background to only around 20%, while two passes results in backgrounds of less than 10% (usually less than 5%). The homopolymer cytidine tails in this batch of pARC5 averaged about 14 residues (Fig. 4B), with the bulk of the sample in the 10-20 residue range. I have noticed somewhat better cloning efficiencies with linker having shorter C-tails (8-10 residues), although well-controlled comparisons have not been done.
20. , 6 including two bandings in CsCI gradients. Plasmids are extracted extensively in CsC1saturated isoamyl alcohol or water-saturated n-butanol in order to remove ethidum bromide. Alkaline lysis methods are avoided because they were found to cause appreciable contamination by small DNA fragments, which interfere with terminal deoxynucleotidyltransferase reactions in later steps. Large-Scale Preparation of Competent Cells Glassware and plasticware are rinsed extensively with distilled water to remove all traces of detergents.