Quantitation of Nucleic Acids and Proteins

  title={Quantitation of Nucleic Acids and Proteins},
  author={Sean R. Gallagher and Philippe R. Desjardins},
  journal={Current Protocols Essential Laboratory Techniques},
Reliable quantitation of nanogram and microgram amounts of DNA and RNA in solution is essential to researchers in molecular biology. Two methods for direct absorbance measurements at 260 nm are described—the first is a traditional cuvette‐based method, and the second is a microvolume method that requires no cuvettes or capillaries. In addition, three fluorescence techniques using Hoechst 33258, ethidium bromide, and PicoGreen reagent are presented in this unit. These five procedures cover a… 

Comparison of Fluorometric Detection Methods for Quantitative Polymerase Chain

The data have indicated the LOD of real‐time PCR method was approximately three orders of magnitude lower than the end‐point measurement method, with a linear range spanning six order of magnitude; 10 fmol to 10 zmol of PCR template.

Optical Design of a Quantitative Microvolume Nucleic Acid Spectrophotometer with Non-Optical Fiber and All Radiation-Hardened Lens Elements

The purity of the nucleic acid samples obtained by extraction/precipitation or adsorption chromatography must be verified with microvolume spectrophotometry to ensure a high success rate of the

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Column Chromatography

This unit is focused on setting up laboratory‐based column chromatography methods using three commonly used chromatographic techniques for separation of proteins, such as size exclusion, ion exchange, and affinity chromatography.

Genotypic diversity and pathogenic potential of Yersinia enterocolitica biotype 2 strains isolated in Brazil

The results of the ERIC-PCR and PFGE showed that the B2strains evaluated in this study had a high genotypic similarity, suggesting that these strains differed little over the 19 year study period and that the environment was a possible source of contamination of humans and animals in Brazil.

Alterations of Genes Involved in Apoptosis and Epigenetic Modulation Associated with Gatifloxacin-Induced Oxidative Stress in Rat Liver

The results showed that the hepatic oxidative stress was associated with increase in the expression of proapoptotic genes and upregulation of genes involved in DNA and histone methylation in gatifloxacin-induced oxidative stress in rat liver.

Chapter 2 Characterization of Phosphorus Forms in Soil Microorganisms

Soil microorganisms act as sink and source of phosphorus (P) and mediate key processes in the soil P cycle, e.g., P mineralization and immobilization (Oberson and Joner 2005). The role of mycorrhizal

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Results confirm gene dispersion and/or a possible contamination of the area with the herbicide, which reinforces global concern of the increase and intensive use of pesticides worldwide.

Recipes for Commonly Encountered Reagents

  • S. Gallagher
  • Biology
    Current Protocols Essential Laboratory Techniques
  • 2018
This unit lists recipes for many of the buffers and other reagents typically found in a molecular biology laboratory, and refers to the unit describing the method.

Assays for determining cell differentiation in biomaterials

This chapter is to give a general overview of the techniques which can be used to determine the differentiation of cells and should act as a guide offering advice as to the selection and optimization of protocols to meet particular needs for cell biomaterials characterization.



Quantitation of DNA and RNA with Absorption and Fluorescence Spectroscopy

This unit describes the traditional absorbance measurement at 260 nm and three more sensitive fluorescence techniques, as well as three microvolume methods that use fiber optic technology in specialized cells or instrumentation to allow quantitation of DNA solutions.

Development and characterization of the NanoOrange protein quantitation assay: a fluorescence-based assay of proteins in solution.

A sensitive fluorescence assay for the quantitation of proteins in solution using the NanoOrange reagent, a merocyanine dye that produces a large increase in fluorescence quantum yield upon interaction with detergent-coated proteins, which is well suited for use with standard fluorescence microplate readers, fluorometers, and some laser scanners.

Spectrophotometric Determination of Protein Concentration

This unit describes spectrophotometric and colorimetric methods for measuring the concentration of a sample protein in solution by comparison with a standard curve or published absorptivity values for that protein.


Effect of pH and ionic strength on the spectrophotometric assessment of nucleic acid purity.

RNA A260/280 ratios are found to be more reliable and reproducible when these spectrophotometric measurements were performed at pH 8.0-8.5 and the ability to detect protein contamination was significantly improved when RNA wasSpectrophotometrically analyzed in an alkaline solution.

Colorimetric protein assay techniques

It is suggested that the most critical issue in the use of a chromogenic protein assay for the characterization of a biopharmaceutical is the selection of a standard for the calibration of the assay; it is crucial that the standard be representative of the sample.