explain:
When using large grinding beads, 2ml u-bottom tube is required, and 1.5ml sharp bottom tube will get stuck;
The size of tissue block should be less than 1-3mm
Before using the beads, it is best to soak them in absolute ethanol for more than 15 minutes, pour out the absolute ethanol and dry them for standby
Several methods to ensure low temperature: A. if possible, the grinding time shall be as short as possible; b. Aluminum alloy pipe adapter (precooling in - 80 ° refrigerator or quick freezing with liquid nitrogen); c. The grinding beads and grinding tubes are kept in a low-temperature refrigerator (- 20 or 4 °) when not in use
DNA extraction
organization Specification and quantity of grinding beads frequency time Recommended solution
plant tissue 2-3 small or 1 large 60Hz 40-90 seconds TPS liquid, not easy to bubble
Heart, kidney and muscle 1 large 65Hz 60 seconds
Stomach, intestine and lung 1 large plus 2 small 65 Hz 90-120 seconds
Liver, brain and spleen 1-2 small 60 Hz 45-60 seconds
blood vessel 1-2 small grains (better effect if proper quartz sand is added) 60 Hz 60 seconds
RNA extraction
organization Specification and quantity of grinding beads frequency time Recommended solution
plant tissue 2-3 small or 1 large 60Hz 40-90 seconds Trizol
Heart, kidney and muscle 1 large 65Hz 60 seconds Trizol
Stomach, intestine and lung 1 large plus 2 small 65 Hz 90-120 seconds Trizol
Liver, brain and spleen 1-2 small 60 Hz 30-60 seconds Trizol
blood vessel 1-2 small grains (better effect if proper quartz sand is added) 60 Hz 60 seconds Trizol
Large 6mm; Medium 4mm; Small 3mm
In order to keep the temperature of the ground sample as low as possible, there are several aspects to pay attention to:
Put the adapter in the - 20 ° refrigerator when not in use; The grinding tube and grinding beads can be placed in a - 80 ° or - 20 ° refrigerator
Under the condition of ensuring the grinding effect, the number of beads should be as small as possible and the grinding time should be as short as possible
Protein extraction
During protein extraction, the best frequency is set at 60Hz, and the grinding time is about 30 seconds longer than that of RNA extraction. The specific grinding time depends on the amount and type of tissue to be ground, such as liver, brain and spleen. Grind with 2 4mm beads for 45 seconds; For those difficult to grind, such as heart, kidney and muscle, use 2ml centrifuge tube, put in 1 large bead of more than 4mm, add 1-2 small beads of 3mm, and grind for 60 seconds. When grinding, add the extract.
Liquid nitrogen grinding
For samples that need special low temperature (enzyme activity test) or are difficult to grind (skin, bone, hair), 2ml tube can be used. Add a 6mm bead. After adding the sample (the sample is cut into small pieces of about 1-2mm), cover the tube tightly, put it into the aluminum alloy adapter, freeze it in liquid nitrogen for about 2 minutes, and grind it at 55 Hz for 15-30 seconds, The grinding effect is almost the same as that of liquid nitrogen. If the grinding effect is not particularly ideal, freeze it again and repeat the grinding. Do not grind for too long, the sample temperature rises and it is easy to tamp.
