The principle of the method is simple; when a pure PCR product of the 16S gene is obtained, sequenced, and aligned against bacterial DNA data base, then the bacterium can be identified. A selected PCR band from each of 40 isolates was sequenced and the bacterium identified to species or genus level using BLAST.

The HIV reverse transcriptase also has ribonuclease activity that degrades the viral RNA during the synthesis of cDNA, as well as DNA-dependent DNA polymerase activity that copies the sense cDNA strand into an antisense DNA to form a double-stranded viral DNA intermediate (vDNA).

Detecting the presence of bacterial DNA by PCR can be useful in diagnosing culture-negative cases of infection, especially in patients with suspected infection and antibiotic therapy.

When identifying bacteria in the laboratory, the following characteristics are used: Gram staining, shape, presence of a capsule, bonding tendency, motility, respiration, growth medium, and whether it is intra- or extracellular.

The methods used for the detection of DNA are:

1. UV- Vis Spectrophotometric analysis.
2. Fluorometric analysis.
3. DNA Precipitation.
4. DNA Gel electrophoresis.
5. Polymerase chain reaction.

Abstract. A molecular hybridization technique using radioactive and non radioactive DNA probes, has been used to detect ASFV DNA immobilized on nitrocellulose paper. It is based on the use of plasmid pRPEL-2 as a hybridization probe.

Two of these mutation detection systems are reviewed here. Both rely on chemical modification of mismatched nucleotides, by either carbodiimide or hydroxylamine and osmium tetroxide. The methods are termed the carbodiimide (CDI) and the Chemical Cleavage of Mismatch (CCM) methods.

Using UV absorbance is one of the most common ways to quantify DNA.

Specifically, the amino acids tyrosine and tryptophan have a very specific absorption at 280 nm, allowing direct A280 measurement of protein concentration. UV absorbance at 280 nm is routinely used to estimate protein concentration in laboratories due to its simplicity, ease of use and affordability.

The A260/A280 provides insight regarding the type of nucleic acid present (dsDNA or RNA) as well as providing a rough indication of purity. In buffered solutions, pure dsDNA has an A260/ A280 of 1.85–1.88 and pure RNA has a ratio of around 2.1.

The Nanodrop does an excellent job at measuring across a wide spectrum that spans UV and visible light. It can’t automatically determine for you that the sample on the pedestal is DNA, RNA or protein – you have to tell the software before beginning measurements so it can report an accurate concentration.

The main advantage the Nanodrop has over the Qubit is the ability to measure the purity of the sample. The 260/280 and 260/230 ratios give an indication of how pure the sample is from protein and salt contaminants. The machine doesn’t require any reagents so sample measurements do not come at a cost.

The main reason people use the Nanodrop is to deduce the purity of their samples. This is generally indicated in two ratios: 260/280 and 260/230. These numbers correspond to the absorbance at the wavelengths 230, 260 and 280 nm.

A 260/280 ratio of ~1.8 is generally accepted as “pure” for DNA; a ratio of ~2.0 is generally accepted as “pure” for RNA. Abnormal 260/280 ratios usually indicate that a sample is contaminated by residual phenol, guanidine, or other reagent used in the extraction protocol, in which case the ratio is normally low.

The 260/230 ratio is used to indicate the presence of unwanted organic compounds such as Trizol, phenol, Guanidine HCL and guanidine thiocyanate. Generally acceptable 260/230 ratios are in the range of 2.0 – 2.2. Values higher than this may indicate contamination with the aforementioned compounds.

To evaluate DNA purity, measure absorbance from 230nm to 320nm to detect other possible contaminants. The most common purity calculation is the ratio of the absorbance at 260nm divided by the reading at 280nm. Good-quality DNA will have an A260/A280 ratio of 1.7–2.0.