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Preparation method and operation flow of RNA agarose gel MOPS(cas 1132-61-2) electrophoresis reagent

1, Material

Total RNA extract. 

2, reagents

(1) 0.1% DEPC water: 200ml double distilled deionized water plus 0.2mlDEPC coke acid, diethyl ester mix, room temperature overnight, autoclave. 
(2) 10x electrophoresis buffer 
MOPS(cas 1132-61-2) 0.4 mol /L (pH 7.0)
NaAc 0.1mol /L
Ethylenediamine tetraacetic acid (EDTA) 10 mmol /L
(3) 50 mL denatured agarose gel (1%)
(4) 10x electrophoresis buffer 5 mL agarose 0.5 g
0.1% DEPC water 36.5 mL
Heated to dissolve, slightly cooled, add 8.5 mL of 37% formaldehyde. 
(5) loading buffer:
50% glycerol, 1 mmmol /LEDTA, 0.4% bromophenol blue, 0.4% xylene blue. 

(6) Formamide (deionized).

3, Apparatus 

(1) Electrophoresis system 

(2) UV perspective instrument

Steps:

Glue:

Weigh 0.5g of agarose powder, add 36.5mL of DEPC water conical flask, heated to completely dissolve the agarose. After cooling slightly, 5 mL of 10x electrophoresis buffer, 8.5 mL of formaldehyde was added. And then in the glue tank filling the gel, insert the comb, the level of use to be used after solidification.

Sample:

The following reagents were mixed in a clean, small centrifuge tube: 2 μl of electrophoresis buffer (10x), 3.5 mL of formaldehyde, 10 mL of formamide, and 3.5 μl of RNA sample. Mix, set 60 ℃ insulation 10min, ice on the cold. Add 3μl of loading buffer to mix, take appropriate amount in the gel sample hole. At the same time point RNA standard sample.

 

 

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