First, the purpose of the experiment
Master the principles and methods of total RNA non-denaturing gel electrophoresis.
Second, the experimental principle
RNA electrophoresis can be carried out under both denaturing and non-denaturing conditions. Non-denaturing electrophoresis uses 1.0%--1.4% gel, and different RNA bands can also be separated, but the molecular weight cannot be judged. The mobility of RNA is linear with the logarithm of the molecular weight only under conditions of complete denaturation. Therefore, when determining the molecular weight of RNA, it is necessary to use a denaturing gel. When it is necessary to quickly detect the integrity of the total RNA sample, a normal 1% agarose gel can be prepared.
Third, experimental materials, utensils and drugs
Mushroom total RNA solution. Electrophoresis, electrophoresis tank, electronic balance, pipette, pipette, microwave oven, UV transmission detector, etc. Agarose, 1XTAE running buffer, 0.5 μg/ml ethidium bromide (EB) 10X loading buffer.
Fourth, the experimental steps
(1) Agarose gel was prepared using 1×TAE running buffer, and 1×TAE running buffer was added to the liquid surface covering gel.
(2) On a clean bench, pipette 4 μl of total RNA sample onto the parafilm. Add 5 μl of 1×TAE running buffer and 1 μl of 10X loading buffer to the bench. After mixing, carefully add the wells.
(3) Turn on the power switch, adjust the voltage to 100V, and make the RNA electrophoresis from the negative electrode to the positive electrode. After about 30 minutes, the gel is placed in the EB dye solution for 5 minutes, and rinsed slightly with water. The results of RNA electrophoresis were observed on a UV transmission detector.
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RNA denatured agarose gel assay
Reagents:
(1) MOPS buffer (10*): 0.4 mol/L morpholinopropanesulfonic acid (MOPS) (Ph7.0), 0.1 mol/L NaAc, 10 mol/L EDTA.
(2) Loading dye: 50% glycerol, 1 mmol/L EDTA, 0.4% bromophenol blue, 0.4% xylene blue.
(3) Formaldehyde.
(4) Deionized formamide. v Electrophoresis tank cleaning: detergent cleaning (usually soaking overnight) - water rinse - ethanol drying - 3% H2O2 filling - room temperature for 10 minutes - 0.1% DEPC water rinse.
operating:
(1) Wash the gelatinous appliance with 70% ethanol and dry it for later use.
(2) Prepare an agarose gel.
1 Weigh 0.5g agarose, put it in a clean 100ml Erlenmeyer flask, add 40ml of distilled water, and heat it in the microwave to make the agarose dissolve completely.
2 To be cooled to 60--70 °C, 9 ml of formaldehyde, 5 ml of 10X MOPS buffer and 0.5 ul of ethidium bromide were added thereto in turn, and uniformly mixed.
3 Fill the agarose gel.
(3) Sample preparation:
1 Take a 500 ul small centrifuge tube treated with DEPC and add the following reagents in sequence: 10 ul MOPS buffer 2 ul, formaldehyde 3.5 ul, formamide (deionized) 10 ul, RNA sample 4.5 ul, and mix.
2 Place the tube in a 60 ° C water bath for 10 minutes, then place on ice for 2 minutes.
3 Add 3 ul of loading dye to the tube and mix.
(4) Loading.
(5) Electrophoresis: 1XMOPS buffer was added to the electrophoresis tank and electrophoresed at a voltage of 7.5 V/ml.
(6) After the electrophoresis is finished, the results are checked under an ultraviolet lamp.
Product Name: Chamomile extract
English Name: Chamomile extract
Plant part used: Flowers
Active ingredient: apigenin
character; Brownish yellow powder
Specification: 10:1
CAS No.: 520-36-5
Efficacy: calm the mind, help digestion, improve skin and resist oxidation.
Function:
(1) Calming effect
The cooling effect of chamomile can treat fever. Its calming and soothing effect can resist allergy, relieve nervous tension, improve insomnia, soothe mental trauma, and improve allergic constitution;
(2) Antispasmodic and analgesic effects
Chamomile has anti spasm and analgesic effects (stomach pain, physiological pain, headache, muscle pain and toothache);
(3) Role of Gynecology
Chamomile can nourish uterus and ovary and improve pelvic inflammation;
(4) Role of digestion
Chamomile can improve digestion, regulate intestines and stomach, and eliminate flatulence;
(5) Role of skin
Chamomile Essential Oil can improve sensitive, dry and peeling skin and increase skin elasticity. It can also treat skin inflammation such as eczema, abscess and acne, promote wound healing, and has excellent effect on regulating sunburned and scalded skin.
Purslane Seed Extract, Chamomile Extract Powder, Morinda Root Extract
Xi'an Tian Guangyuan Biotech Co., Ltd. , https://www.tgybiotech.com