Biotech Lab Protocols, Tips & Tricks

PURPOSE: To perform ligation of given plasmid DNA(vector,insert) to use it for transformation.

REAGENTS: Plasmid DNA fragments , T4 DNA ligase , Ligase buffer , Wash buffer and Elution buffer.

INSTRUMENTS: Microcentrifuge, Heating block.

PRINCIPLE: DNA ligation is the process of joining together two DNA molecule ends (either from the same or different molecules). This process is accomplished both invitro and invivo by the enzyme DNA Ligase. It involves

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PURPOSE: To Purify DNA by High Speed Centrifugation of Agarose Gel Slices.

REQUIREMENTS: T.E buffer , ddH2O,DNA Trans-illuminator, High Speed Centrifugation, Long wavelength U.V light.

PRINCIPLE: High speed centrifugation: With high speed centrifugation, the agarose matrix is compressed, and/or

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PURPOSE: To digest given DNA sample using restriction enzyme EcoR1.

REQUIREMENTS: DNA ,Distilled water, Restriction enzyme ,Enzyme buffer, Heating block, Electrophoresis assembly

PRINCIPLE: A Restriction digest is a procedure to prepare DNA for analysis or other processing. The enzymatic technique can be used for cleaving DNA molecules at specific sites ,

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PURPOSE: To determine the DNA concentration by spectrophotometric estimation.

PRINCIPLE: The easiest way of determinig DNA concentration is through spectrophotometric analysis.Since nitrogenous bases absorb UV light,the more concentrated the DNA solution, the more UV light it will absorb. The concentration of pure double stranded DNA

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PURPOSE: To determine the presence or absence of PCR products and quantify the size (length of the DNA molecule) of the product.

MATERIALS REQUIRED: Agarose, TBE Buffer, 6X Sample Loading Buffer. DNA ladder standard,Electrophoresis chamber, Power supply, Gel casting tray and combs, Glove,6X DNA Loading Dye - 10 ml. , Pipette and tips

PRINCIPLE:

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