Dna Isolation Mol Bio

a procedure used to isolate DNA from the nucleus of cells from other cellular components

Process of purification of DNA from sample using a combination of physical and chemical methods

DNA extraction is a Process of purification of DNA from sample using a combination of

Methods used to isolate DNA are dependent on the source, age and size of the sample

DNA extraction is used to isolate DNA. And are dependent on the - - - - sample

Example of DNA extraction

obtain DNA in a relatively purified form which can be used for further investigations

DNA can be sourced in any

Common sources for DNA isolation:

Isolation of DNA basically consists of

4 major sites of DNA extraction

To extract DNA from a tissue/cells of interest, the cells have to be separated and the cell membranes have to be disrupted

Extraction buffers

The “Extraction buffer” , EDTA and SDS helps in carrying out these processes.

The - - - of preparation of Cell extract , EDTA and SDS helps in carrying out these processes.

Having lysed the cells, the final step in the preparation of a cell extract is

In addition to the DNA the cell extract will contain significant quantities of detergents, proteins, salts and reagents used during cell lysis step and RNA

Other substances found in the Cell extract aside from DNA

variety of procedures can be used to remove these contaminants, leaving the DNA in a pure form.

Cell debris and partially digested organelles can be pelleted by centrifugation leaving the cell extract as a reasonably clear supernatant.

can be pelleted by centrifugation leaving the cell extract as a reasonably clear supernatant.

Cell debris and partially digested organelles can be pelleted by centrifugation leaving the cell extract as a reasonably clear

Most commonly used procedure in dna extraction

Organic Extraction Reagents

Non-ionic detergent, salt, buffer, EDTA designed to lyse outer cell membrane, but will not break down nuclear membrane

is a chelating agent of divalent cations such as Mg 2+

removal of protein by digesting with proteolytic enzymes

remove proteinaceous material

keep the solution at defined pH; ensure stability of DNA and long term storage

Help Precipitation of DNA

Will lyse the cell membrane

The DNA released in the solution is extracted with

DNA is precipitated from the aqueouslayer by the addition of ice cold

Precipitated DNA is washed with

After the DNA is washed it is dried under and resuspend in

Organic extraction

Does not use organic reagents such as phenol or chloroform

In inorganic DNA Extraction. Digested proteins are removed by

Alteration of DNA mobility

However, if salts are not adequately removed, problems could occur with the procedure due

To remove proteinaceous material in inorganic extraction. What is added in the incubated Ice

In inorganic extraction DNA is precipitated by theaddition ofroomtemperature

Relies on the fact that the nucleic acids maybind (adsorption) to the solid phase (silica orother) depending on the pH and the saltconcentration of the buffer

spin column using a silica-based extractionmethod is used

Nucleic acids are attracted to the silica bead under high chaotropic salt concentrations

Nucleic acids are attracted to the - - - - under high chaotropic salt concentrations

Nucleic acids are attracted to the silica bead under high--- concentrations

In minicolumn extraction how many times is it washed

Steps in minicolumn extraction

The Cells of a sample is broken with a

Elution buffer of minicolumn extraction

In minicolumn extraction it removes the nucleic acid from the membrane and the nucleic acid is collected from the bottom of the column

Degrades single stranded RNA

Dissolves RNAse

Lyse Gram-negative bacterial cell wall

Lyse Gram-positive bacterial cell wall

Solubilize cell membrane lipids

Separates DNA from other cellularmaterials

Phenol Chloroform is also called as

Precipitates DNA from solution

Dissolves precipitated and dried DNA

The most frequently used method of concentration is

With a concentrates solution of DNA one can use a---__to pull out the adhering DNA strands

For dilute solutions precipitated DNA can be collectedby - - - - and redissolving in an appropriatevolume of water

For - - - _precipitated DNA can be collected by centrifugation and redissolving in an appropriate volume of water

DNA concentrations can be accurately measured by

The amount of UV radiation absorbed by a solution of DNA is directly proportional to the

With a pure sample of DNA the ratio of theabsorbancies at 260 nm and 280 nm (A260/A280) is