Journal: PLoS ONE

Article Title: Fidelity Index Determination of DNA Methyltransferases

doi: 10.1371/journal.pone.0063866

Figure Lengend Snippet: Radioactive methylation assay using E. coli DNA-comparison between restriction enzyme digested substrate versus undigested substrate. The dilution series and presentation of data are the same as in Figure 3 . A) H 3 -methyl incorporation by M.EcoKDam. At a dilution of 1/2 4 , the substrate is fully methylated and after a dilution of 1/2 2 , the CPMs start to increase indicating star activity. This results in an FI of 4. B) H 3 -methyl incorporation by M.EcoKDam with DNA that has been digested with MboI restriction enzyme to remove all M.EcoKDam cognate sites. After a dilution of 1/2 2 , the CPMs start to increase, indicating methylation at non-cognate sites. C) H 3 -methyl incorporation by M.HhaI. At a dilution of 1/2 8 , the substrate is fully methylated and after a dilution of 1/2 3 , the CPMs start to increase indicating star activity. This results in an FI of 32. D) H 3 -methyl incorporation by M.HhaI with DNA that has been digested with HhaI restriction enzyme to remove all M.HhaI cognate sites. After a dilution of 1/2 3 , the CPMs start to increase, indicating methylation at non-cognate sites.

Article Snippet: Digestion of cognate sites with restriction enzyme The digestion reaction contained 23 µg of E. coli genomic DNA pre-sheared to 1 kB fragments by a Covaris s-series sonicator, and 200 units of either MboI (NEB #R0147), HhaI (NEB #R0139) or AluI (NEB #R0137) restriction enzyme in NEB4.

Techniques: Methylation, Activity Assay

Journal: Environmental Microbiology

Article Title: Use of stable isotope-labelled cells to identify active grazers of picocyanobacteria in ocean surface waters

doi: 10.1111/j.1462-2920.2008.01793.x

Figure Lengend Snippet: Self-organizing tree (SOTA) of terminal restriction fragment length polymorphism (T-RFLP) profiles emerging from the 18S rRNA sequences from the different experimental treatments. Samples digested (A) with HhaI and (B) with RsaI. Time 0: T-RFLP profile of rRNA from the entire eukaryotic community at the beginning of the experiment. Unlabelled Pro and Unlabelled Syn: T-RFLP profiles of the unlabelled eukaryotic rRNA, collected from the density gradient, from the bottles that were incubated with labelled Prochlorococcus and Synechococcus respectively. Labelled Pro and Labelled Syn : T-RFLP profiles of the heavily labelled eukaryotic rRNA that was collected from the same gradient. A and B next to the data points represent the two biological replicates.

Article Snippet: PCR products were digested with the restriction endonucleases HhaI and RsaI (New England Biolabs, Ipswich, MA, USA).

Techniques: Terminal Restriction Fragment Length Polymorphism, Incubation