Ecorii

Manufactured by Thermo Fisher Scientific

Sourced in Lithuania

EcoRII is a type II restriction endonuclease enzyme isolated from the bacterium Escherichia coli. It recognizes and cleaves the palindromic DNA sequence 5'-CCWGG-3'. EcoRII is commonly used in molecular biology applications such as DNA digestion, fragment analysis, and genetic engineering.

Automatically generated - may contain errors

Lab products found in correlation

8 protocols using ecorii

Phage DNA Extraction and Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

Aliquots of phage suspension (10¹¹–10¹² PFU/ml) were subjected to phenol/chloroform extraction and ethanol precipitation as described by Carlson and Miller [38] . Isolated phage DNA was subsequently used in restriction analysis, for PCR or was subjected to genome sequencing. Restriction digestion was performed with BamHI, EcoRI, EcoRII, EcoRV, HindIII, KpnI, MboI, NheI, NotI, PstI, PvuI, SnaBI, SspI, VspI and XbaI restriction endonucleases (Fermentas) according to the supplier's recommendations. DNA fragments were separated by electrophoresis in a 0.8% agarose gel containing ethidium bromide. Restriction analysis was performed in triplicate to confirm the results.

Šimoliūnas E., Kaliniene L., Stasilo M., Truncaitė L., Zajančkauskaitė A., Staniulis J., Nainys J., Kaupinis A., Valius M, & Meškys R. (2014). Isolation and Characterization of vB_ArS-ArV2 – First Arthrobacter sp. Infecting Bacteriophage with Completely Sequenced Genome. PLoS ONE, 9(10), e111230.

+ Open protocol

+ Expand

Genomic DNA Methylation Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

We extracted the genomic DNA from 10-day-old seedlings. A total of 10 μg of genomic DNA was digested with the methylation sensitive restriction enzymes, 20 U of MspI, HpaII, BstNI and EcoRII (Fermentas, Thermo Scientific) and then separated by electrophoresis in a 0.8% agarose gel. We blotted the digested and separated genomic DNA onto a nylon membrane and analyzed by Southern blot analysis using ³²P-labeled [TTTAGGG]₇₀ oligonucleotides as a probe. Hybridization and the washing of the membranes were carried out according to the method described in TRF assay.

Lee W.K, & Cho M.H. (2016). Telomere-binding protein regulates the chromosome ends through the interaction with histone deacetylases in Arabidopsis thaliana. Nucleic Acids Research, 44(10), 4610-4624.

+ Open protocol

+ Expand

Phage DNA Extraction and Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

The aliquot of high-titer (10¹¹–10¹² PFU/mL) phage suspension was subjected to phenol/chloroform extraction and ethanol precipitation, as described by Carlson and Miller [33 ]. The isolated phage DNA was subsequently used for PCR and restriction analysis, or it was subjected to genome sequencing. The restriction digestion was performed with Bsu15I, Csp6I, DraI, EcoRII, EcoRV, HhaI, MboI and NdeI restriction endonucleases (Thermo Fisher Scientific, Vilnius, Lithuania), according to the supplier’s recommendations. The DNA fragments were separated by electrophoresis in a 0.8% agarose gel containing ethidium bromide. A restriction analysis was performed in triplicate to confirm the results.

Žukauskienė E., Šimoliūnienė M., Truncaitė L., Skapas M., Kaupinis A., Valius M., Meškys R, & Šimoliūnas E. (2021). Pantoea Bacteriophage vB_PagS_AAS23: A Singleton of the Genus Sauletekiovirus. Microorganisms, 9(3), 668.

+ Open protocol

+ Expand

Phage DNA Extraction and Restriction Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

The aliquots of high-titer (10¹¹–10¹² PFU/mL) phage suspension were subjected to phenol/chloroform extraction and ethanol precipitation, as described by Carlson and Miller [84 ]. The isolated phage DNA was subsequently used in restriction analysis, for PCR, or it was subjected to genome sequencing. The restriction digestion was performed with Bsu15I, Csp6I, DraI, EcoRII, EcoRV, HhaI, MboI, and NdeI restriction endonucleases (Thermo Fisher Scientific, Vilnius, Lithuania), according to the supplier’s recommendations. The DNA fragments were separated by electrophoresis in a 0.8% agarose gel containing ethidium bromide. The restriction analysis was performed in triplicate and Figure 4 shows a representative result.

Šimoliūnienė M., Žukauskienė E., Truncaitė L., Cui L., Hutinet G., Kazlauskas D., Kaupinis A., Skapas M., de Crécy-Lagard V., Dedon P.C., Valius M., Meškys R, & Šimoliūnas E. (2021). Pantoea Bacteriophage vB_PagS_MED16—A Siphovirus Containing a 2′-Deoxy-7-amido-7-deazaguanosine-Modified DNA. International Journal of Molecular Sciences, 22(14), 7333.

+ Open protocol

+ Expand

Phage DNA Extraction and Restriction Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

The aliquots of phage suspension (10¹¹–10¹² PFU/mL) were subjected to phenol/chloroform extraction and ethanol precipitation, as described by Carlson and Miller [22 ]. The isolated phage DNA was subsequently used in restriction analysis, for PCR, or it was subjected to genome sequencing. The restriction digestion was performed with BamHI, Bsu15I, Csp6I, DraI, EcoRII, Eco32I, HhaI, MboI, MfeI, NdeI, and RsaI restriction endonucleases (Thermo Fisher Scientific, Vilnius, Lithuania), according to the supplier’s recommendations. The DNA fragments were separated by electrophoresis in a 0.8% agarose gel containing ethidium bromide. A restriction analysis was performed in triplicate to confirm the results.

Šimoliūnas E., Šimoliūnienė M., Kaliniene L., Zajančkauskaitė A., Skapas M., Meškys R., Kaupinis A., Valius M, & Truncaitė L. (2018). Pantoea Bacteriophage vB_PagS_Vid5: A Low-Temperature Siphovirus That Harbors a Cluster of Genes Involved in the Biosynthesis of Archaeosine. Viruses, 10(11), 583.

+ Open protocol

+ Expand

Phage DNA Characterization by Enzymatic Assays

Check if the same lab product or an alternative is used in the 5 most similar protocols

Aliquots (100–150 μl) of phage suspensions (10¹¹–10¹² PFU/ml) were subjected to phenol/chloroform extraction and ethanol precipitation. Isolated phage DNA was used for restriction analysis with Eco32I, MboI, EcoRII, SalI and Csp6I restriction endonucleases (Thermo Fisher Scientific) according to supplier’s recommendations. In vitro DNA glycosylation tests were performed in the Epi Buffer using T4 phage β-glucosyltransferase (T4 BGT) and UDP-glucose from the EpiJET 5-hmC and 5-mC Analysis Kit (Thermo Fisher Scientific). DNA fragments were separated by electrophoresis in a 0.8% agarose gels stained with ethidium bromide.

Gordeeva J., Morozova N., Sierro N., Isaev A., Sinkunas T., Tsvetkova K., Matlashov M., Truncaitė L., Morgan R.D., Ivanov N.V., Siksnys V., Zeng L, & Severinov K. (2018). BREX system of Escherichia coli distinguishes self from non-self by methylation of a specific DNA site. Nucleic Acids Research, 47(1), 253-265.

+ Open protocol

+ Expand

Quantitative Methylation Assay for M.MpeI and Dcm

Check if the same lab product or an alternative is used in the 5 most similar protocols

3-fold serial dilutions of M.MpeI (0.78 μM – 3.2 nM) were incubated with 160 μM SAM or CxSAM substrate and pUC19 plasmid DNA (100 ng) for 4 hrs at 37°C in M.MpeI reaction buffer (10 mM Tris Cl, 50 mM NaCl, 1 mM DTT, 1 mM EDTA, pH 7.9 at 25°C) in a 5 μL volume. 2.5 μL of the reaction was then incubated with the appropriate restriction enzyme, and the plasmid DNA was simultaneously linearized with HindIII-HF (NEB) in a final digestion volume of 25 μL. HpaII (NEB) recognizes CCGGs (13 sites). Samples were treated with 1 μL Proteinase K at 37°C for 10 min, separated on 1% TAE Agarose gel, and visualized with SYBR Safe DNA Gel Stain (Thermo-Fisher).
A similar assay was performed with the Dcm proteins except that serial 3-fold dilutions encompassed 0.26 μM – 3.2 nM and activity was instead assessed with EcoRII (Thermo), which recognizes CCWGGs (7 sites). The plasmid was also linearized with NdeI (NEB).

Wang T, & Kohli R.M. (2020). Discovery of an unnatural DNA modification derived from a natural secondary metabolite. Cell chemical biology, 28(1), 97-104.e4.

+ Open protocol

+ Expand

Methylation Analysis of Putative DNA MTases

Check if the same lab product or an alternative is used in the 5 most similar protocols

To assess the methylation ability of the putative DNA MTases, Xe(EV), Xe(XvDMT2), and Xe(XvDMT2) were grown in XVM2 medium to an OD₆₀₀ of 0.6, and bacterial cells were harvested using centrifugation. Genomic DNA was extracted using a genomic DNA extraction kit (Qiagen). The DNA quantities and quality were checked by NanoDrop One (Thermo Scientific). The genomic DNA (1000 ng) was digested for 15 min using 2.5 units of MspJI (NEB, Ipswich, MA, USA), McrBC (NEB), EcoRII (Thermo Scientific), and Bsp119I (Thermo Scientific) at the optimal incubation temperature for each enzyme. The digested genomic DNA was electrophoresed in a 1 % agarose gel at 100 V for 90 min.

Park H.J., Seong H.J., Lee J., Heo L., Sul W.J, & Han S.W. (2021). Two DNA Methyltransferases for Site-Specific 6mA and 5mC DNA Modification in Xanthomonas euvesicatoria. Frontiers in Plant Science, 12, 621466.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!