Category: Lab Notes: Recombinant DNA Technology
KOD DNA Polymerase
Also known as KOD HiFi DNA Polymerase High-fidelity thermostable DNA polymerase for error-free polymerase chain reaction. Originally isolated from Thermococcus…
Phusion DNA Polymerase
Overview: High-fidelity thermostable DNA polymerase for error-free polymerase chain reaction. Possess 3′ to 5′ exonuclease activity for proofreading activity Produces…
Principle of Alkaline Lysis Method
The alkaline lysis method selectively purifies plasmid DNA from other cellular components of the bacterial cells including chromosomal DNA. Controlled…
Plasmid Isolation by Alkaline Lysis Method
Alkaline lysis method of plasmid isolation was originally developed by Birnboim & Doly (1979). In this procedure, bacteria harbouring the desired…
Protocol – Growing Large Volume of Liquid Culture of Escherichia coli for Large-Scale Plasmid Isolation
Large-scale isolation of plasmid requires a large volume of E. coli culture. Large scale plasmid isolation procedures are termed, midiprep (25 – 50 ml starting culture volume) and maxiprep (100 – 500 ml starting culture volume). A starter culture is initially prepared by inoculating a colony in a small volume (2 – 10 ml) of culture medium. Large culture volume is prepared by diluting starter culture in a ratio of 1: 100 to 1: 1000 in the growth medium.
Migration of Bromophenol Blue and Xylene Cyanol in Agarose Gel Running in TBE or TAE Electrophoresis Buffer
Bromophenol blue and Xylene cyanol are the two most commonly used tracking dyes for the analysis of DNA on agarose gel electrophoresis. These negatively charged dyes not only help in monitoring the progress of agarose gel electrophoresis but also allow easy monitoring of sample loading process onto the wells of agarose gel. Their position in relation to DNA fragments is an important information that helps to determine how far a gel must be run without accidentally letting the DNA fragments of interest run out of the gel and at the same time ensuring good resolution among different size DNA fragments.
Recommended Antibiotic Concentrations for the Selection and Growth of Escherichia coli Carrying a Plasmid
Most of the commonly used plasmid vectors carry at least one antibiotic resistance gene that allows plasmid-containing bacterial cells to…
Protocol – RNA Isolation from Tissue Using Trizol
Overview: ♦ Tissues are composed of multiple cell layers that are tightly glued together by extracellular matrix. ♦ Solubilizing all…
Protocol – RNA Isolation from Suspension Cell Culture Using Trizol
Overview: Cell lines are a very useful tool for life science research. Isolation of total RNA from cell lines is…
Protocol – RNA Isolation from Escherichia coli Using Trizol
Overview: Escherichia coli (E. coli) is one of the well characterized bacteria and is extensively used in both applied and…
Protocol – RNA Isolation from Adherent Monolayer Cell Culture Using Trizol
Overview: Cell lines are very useful tools for life science research. Isolation of total RNA from cell lines is often…
List of Monophasic Solution for RNA Extraction
TRIzol ™ Reagent (Invitrogen) Supplied by ThermoFisher Scientific TRI Reagent® Supplied by MRC, Sigma Aldrich, Zymoresearch RNAzol® (Tel-Test) RNA Stat-60…
Isolation of RNA Using Trizol
Trizol is a very popular reagent among researchers for RNA isolation due to the simplicity and reliability of the isolation…
Protocol – Preparing RNA free of DNA Using DNase I Digestion
Overview DNase I is an endonuclease that selectively cleaves phosphodiester bonds of both single- and double-stranded DNA without cleaving RNA….
Preparing RNA free of DNA
DNA-free RNA preparations are absolutely required for applications like gene expression analysis using PCR-based methods (e.g., real-time PCR). No RNA…