RNA isolation / purification Bacteria Gram negative

RNA quantification for appropriate concentration and quality (260/280 ratio) is an important step before downstream analysis (including sequencing, RT-qPCR, etc.). Having insufficient RNA quantities or a high salt or phenol in the RNA product can lead to variable or irreproducible downstream results. The various methods used for RNA quantification include: 1. UV spectrophotometric (challenges include: low sensitivity, cannot distinguish between nucleic acid species), 2. Fluorescence-based (challenges include: requires standards, cannot measure amplifiability, not sequence-specific), and 3. RT-PCR (challenges include: requires standards, time-intensive, costly). In order to overcome these challenges, and also to ensure the proper quantification and quality control for RNA product, it is important to use at least two or more methods in order to discard any inconsistencies. Using standards for calibrations increases the sensitivity range for RNA detention (fluorescence- and RT-PCR-based methods). When using RT- PCR, it is important to choose correct primers, aligning to the desired site on the template and of appropriate product length, along with positive, negative and loading controls. It is also important to have at least two primer pairs in order to confirm results.

RNA quantification for appropriate concentration and quality (260/280 ratio) is an important step before downstream analysis (including sequencing, RT-qPCR, etc.). Having insufficient RNA quantities or a high salt or phenol in the RNA product can lead to variable or irreproducible downstream results. The various methods used for RNA quantification include: 1. UV spectrophotometric (challenges include: low sensitivity, cannot distinguish between nucleic acid species), 2. Fluorescence-based (challenges include: requires standards, cannot measure amplifiability, not sequence-specific), and 3. RT-PCR (challenges include: requires standards, time-intensive, costly). In order to overcome these challenges, and also to ensure the proper quantification and quality control for RNA product, it is important to use at least two or more methods in order to discard any inconsistencies. Using standards for calibrations increases the sensitivity range for RNA detention (fluorescence- and RT-PCR-based methods). When using RT- PCR, it is important to choose correct primers, aligning to the desired site on the template and of appropriate product length, along with positive, negative and loading controls. It is also important to have at least two primer pairs in order to confirm results.

Get tips on using Wizard® Plus Midipreps DNA Purification System Technical Bulletin to perform Plasmid Isolation Proteus mirabilis

Get tips on using Wizard® Plus SV Minipreps DNA Purification System Technical Bulletin to perform Plasmid Isolation Streptomyces spp

Get tips on using Lipofectamine® RNAiMAX Transfection Reagent to perform siRNA / miRNA gene silencing Human - siRNA negative control Lipid

Get tips on using EndoFree Plasmid Mega Kit (5) to perform DNA isolation / purification Plasmid purification

Get tips on using PhosphoProtein Purification Kit (6) to perform Protein tag Purification of phosphorylated proteins

Get tips on using QIAGEN Plasmid Plus Midi Kit (25) to perform DNA isolation / purification Plasmid purification

Get tips on using MinElute PCR Purification Kit to perform DNA gel extraction / PCR product purification Product size < 15Kb

Get tips on using Purelink PCR Purification Kit to perform DNA gel extraction / PCR product purification Product size < 15Kb