Tuesday, March 31, 2009

Protein Assay using UV-Spectrophotometer at 280 nm

It is possible to estimate protein concentration in a solution by using simple spectrometer. Absorption of radiation in the near UV (280 nm) by proteins depends on the Tyrosine and Tryptophan content (also to a very small extent on the amount of Phenylalanine and disulfide bond).

Sunday, March 29, 2009

Surfactant Properties: How to Quantitatively Measure

In order to measure the properties of surfactant, we can use the surface tension, emulsification activity, and hemolytic activity as the parameters. Here are the methods:

Tuesday, March 24, 2009

DNA Extraction Using Prepman Ultra

In my previous posting I’ve slightly given the bacterial DNA extraction method using Instagene matrix. In this particular posting, I want to give you the more simple method to extract DNA, which is using Prepman Ultra. According to the producer, Applied Biosystem, Prepman ultra is applicable to successfully preparing DNA template from bacteria, yeast, filamentous fungi, both from a plate or from tissue smears.

PCR Components

The Polymerase Chain Reaction (PCR) created a revolution in molecular biology research and its applications. PCR is an in vitro method that enzymatically amplifies specific DNA sequences using oligonucleotide primers that flank the region of interest in the target DNA. The principle involves a repetitive series of cycles each of which consist of template denaturation, primer annealing, and extension of the annealed primers by a DNA polymerase to create the exponential accumulation of a specific fragment whose ends are determined by the 5’ ends of the primers. The PCR is so named because it involves a polymerase and the products synthesized in each cycle can serve as templates in the next so the number of DNA copies approximately doubles at every cycle to create a chain reaction similar to the principles in a nuclear reactor.

Sunday, March 22, 2009

Bradford Method: Colorimetric Protein Assay

Bradford method is a common colorimetric method to determine protein concentration in a sample solution. The Bradford method of protein determination is based on the binding of a dye, Coomasie Blue G, to the protein. This binding shifts the absorbtion maximum of the dye from red to blue. The absorbance of the solution is measured at 595 nm and is proportional to protein concentration when compared to a standard curve. Two types of assay are described here: the standard assay, which is suitable for measuring between 10 and 100 microgram of protein, and the microassay, which detects between 1 and 10 microgram of protein.

Friday, March 20, 2009

Quantitative Estimation of DNA Concentrations

DNA, RNA, and protein strongly absorb ultraviolet light in the 260 to 280 nm range. UV spectroscopy can be used as a quantitative technique to measure nucleic acid concentration and protein contamination. Nucleic acids strongly absorb at 260 nm and less strongly at 280 nm while proteins do the opposite. The general rules for determining the concentrations of nucleic acids at 260 nm are:

Thursday, March 19, 2009

Estimation of DNA Concentrations

The determination of the concentration of DNA or RNA in solution is a fundamental task in molecular biology. DNA is usually the limiting reagent in most experiments; therefore, the knowledge of its concentration is critical. Determination of the DNA concentration can be estimated either by qualitatively comparing the fluorescence of DNA bands in an agarose gel to a standard or by spectrophotometric means.

Tuesday, March 17, 2009

Measurement of Biosurfactant Activity

Biosurfactant activity was measured by an oil displacement test. This method is so sensitive that only a small amount of sample is required to measure the surfactant activity. Here is the method:

Wednesday, March 11, 2009

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