DNA, purified

A. sample of DNA purified from MycobacUrium tubercvioiis contain.s 15.1% adenine on a molar basis. What are the percentages of the other bases present ... 

The second approach is not so limited by population complexity or information content. Here 16S rRNA genes are "shotgun cloned" using DNA purified from available biomass (7,8). In this case, the originating population complexity does not matter since the rRNA genes are clonally isolated from a recombinant library of DNA directly derived from the natural microbial community. The different clones are sorted and sequenced using rapid sequencing techniques. 

Table II DNA-binding studies on furazolidone Pig hepatocytes were incubated with 50 nM C-furazolidone for 16 hr (starting 26 hr after isolation), or C-leucine for 40 hr (starting 4 hr after isolation). Cells were homogenized, the chromatin collected by centrifugation and DNA purified by phenolic extraction...

Fig. 3 Appearance of fragments protected from micrococcal nuclease digestion following incubation of Xenopus sperm chromatin in homologous egg extract. Sperm nuclei were used without extract incubation (lane 2) or following incubation of 50 ng DNAZ/il egg LSS for 3 min (lane 3), 10 min (lane 4), 30 rain (lane 5), or 120 min (lane 6). Samples were diluted and the sperm nuclei were isolated by centrifugation. The samples were digested with micrococcal nuclease for 1 min and the DNA purified and separated by a 1.4% Tris-borate-EDTA agarose gel. Lane I shows molecular weight markers in lOO-base-pair increments.

To purify the genomic DNA fragments cross-linked into archaeal nucleosomes, the MN digestion is stopped as described above, and the lysate then treated with proteinase K (3(X) p-g/ml) for 3-4 hr at 37°. The formaldehyde cross-links are reversed by incubation at 65° for 6 and the nucleosomal DNA purified by phenol/chloroform extraction, chloroform extraction, and ethanol precipitation. Exposure to RNase A (40 p-g/ml) for 30 min at 37°C yields a solution containing only 60 bp nucleosomal DNA fragments. 

Digest the PCR product and the destination vector pMS 101C with the BssHII and Sa/I-HF restriction enzymes. The amount of each reagent has to be adapted to the amount of DNA purified in step 2. For 10 pg of DNA, prepare the following solutions ... 

Process the two digestion reactions through DNA Clean Concentrator-25 columns (two columns for each reaction), following the manufacturer s instructions. Wash the columns twice, with 300 and 400 lL of DNA Wash Buffer, respectively, and elute with 35 pL of H2O prewarmed at 65 °C. Pool the eluates to obtain ca. 70 pL of purified DNA for the insert and for the vector. Examine 2 pL on a DNA agarose gel to ensure that single bands of the correct size were obtained. Determine the total amount of DNA purified by measuring DNA concentration. 

Figure 5.4 IR-MALDI mass spectrum of large DNA analyzed from glycerol as a matrix. The spectrum depicts single-stranded DNA molecules of length up to 1.4 kb generated from restriction-digested plasmid DNA purified by ethanol precipitation. The...

Preliminary results indicate that the DNA purified after micrococcal nuclease digestion of the enzyme—DNA complex activates more efficiently the DNA-free poly(ADP-ribose) polymerase than the total purified sDNA and much more than the purified nucleosomal core particles DNA (data not shown). Ohgushi et al. [8] and Benjamin and Gill [7] correlate the activation of poly(ADP-ribose) polymerase only to its binding to nicks or ends and not to a specific DNA sequence. Although our observations of poly(ADP-ribose) polymerase-sDNA complexes by electron microscopy and by polyacrylamide gel analysis do not exclude the possibUity that the enzyme is activated preferentially by internal nicks on sDNA fragments, they raise the question whether... 

Plasmid DNA Purify plasmid DNAs by equilibrium centrifugation in cesium chloride containing ethidium bromide. Dissolve the preparations in 10 mM Tris-HCl, pH 8.0, 0.1 mM EDTA (final concentration of DNA was 2 mg/ ml), and store at —20°C. 

Virus DNA Purified AcNPV./flcZ (also called AcRP23./recombinant virus plaques (Kitts et al., 1990). Use of purified BACPAK6 or BaculoGold DNA will yield virtually 100% recombinant virus plaques (Kitts and Possee, 1993). 

Use each time one-fourth of the PCR mixture from the previous cycle for the next round of binding, selection, and amplification To avoid formation of artifactual DNA, purify the PCR products obtained after every other round on an 8% nondenaturing PAGE (essentially as described for the randomer purification but in the absence of urea)... 

In research conducted by Sabbioni and Girardi, NAA was developed for the simultaneous analysis of heavy metals in microsamples of potential metal binding components such as metalloenzymes (calf intestine alkaline phosphatase, cow milk xanthine oxidase, calf thymus deoxynucleotidyltransferase) and nucleic acid (calf thymus DNA), purified by size-exclusion chromatography and ion-exchange chromatography. 

Even without addition of RNA-polymerase, DNA purified from histone possessed considerably (five times) more activity as regards RNA synthesis on account of the pol3onerase still remaining in it (Huang and Bonner, 1962). These workers concluded from their findings that histones repress mRNA synthesis on DNA. 

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