PCR Hot start PCR

Get tips on using Ni-NTA Fast Start Kit (6) to perform Protein tag Purification of His-tagged proteins

Get tips on using Quick Start™ Bradford Protein Assay Kit 1 to perform Protein quantification Mammalian cells - BV-2

Get tips on using peqGOLD Cycle‐Pure Kit to perform DNA gel extraction / PCR product purification Product size > 15Kb

Get tips on using QIAquick Gel Extraction Kit to perform DNA gel extraction / PCR product purification Product size < 15Kb

Get tips on using MinElute Reaction Cleanup Kit (250) to perform DNA gel extraction / PCR product purification Product size < 15Kb

Get tips on using MinElute Gel Extraction Kit (250) to perform DNA gel extraction / PCR product purification Product size < 15Kb

Get tips on using QIAEX II Gel Extraction Kit (150) to perform DNA gel extraction / PCR product purification Product size > 15Kb

Get tips on using Zymoclean™ Gel DNA Recovery Kit to perform DNA gel extraction / PCR product purification Product size > 15Kb

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.