1xbet best casino website

MONOCOT 1xbet best casino website ISOLATION

1. Collect 1xbet best casino website tissue and store at -20°C.
2. Grind leaf tissue to a fine powder with 1xbet best casino website in a prechilled mortar [Store mortar and pestle at -20°C or -80°C].
3. Transfer ground tissue with a chilled paintbrush into a well-chilled, 50-1xbet best casino website polypropylene tube and store at -20°C.Do not 1xbet best casino website powder thaw.All tools should be chilled in 1xbet best casino website .
4. Add sodium bisulfite to the extraction buffer and adjust pH with 5N NaOH to 7.8–8.0. Heat the extraction buffer to 65°C and add 15–20 1xbet best casino website to 10–15 1xbet best casino website of frozen tissue. Mix well.

1. Incubate in a 65°C water bath for 30 min; invert the tubes every 5–10 1xbet best casino website .
2. Add chloroform:isoamyl alcohol [24:1 v/v] to the top of the 1xbet best casino website and mix vigorously.
3. Centrifuge 15 min at 4,500 rpm. Transfer upper phase into a new 50-1xbet best casino website tube by pouring through 2–4 layers of cheesecloth or pipette off the upper phase if the interphase does not appear solid.
4. Add 2 volumes (fill tube to top) of cold [-20°C] 95% EtOH and mix gently to precipitate the 1xbet best casino website . Place at -20°C for 30–60 min.
5. Pour off the 95% EtOH, wash with fresh 70% EtOH, and add 30 ml of cold 70% EtOH. Mix gently for a few minutes. 1xbet best casino website can be left indefinitely in 70% EtOH [can leave overnight at -20°C].
6. Rewash with fresh 70% EtOH if the 1xbet best casino website is still discolored. Remove 1xbet best casino website by hooking it on a Pasteur pipette and blot off excess 70% EtOH with a Kimwipe. Transfer 1xbet best casino website to the bottom of a sterile 1.5-ml microfuge tube. If making many extractions, centrifuge wet 1xbet best casino website gently, then invert tube a few minutes to let the liquid drain out. Dry 1xbet best casino website at RT for 2–3 hours.
7. Dissolve 1xbet best casino website in1xbet best casino website TE [0.5–1 mL depending on pellet size]. Incubate in a 65°C water bath until dissolved with gentle inversion every 30 to 60 minutes or until 1xbet best casino website is dissolved [may take several hours].
8. Centrifuge 10 1xbet best casino website in microcentrifuge at 13,000 rpm to remove any material that does not go into solution.
9. Determine the 1xbet best casino website concentration by separating a 1:20 dilution [1 1xbet best casino website :20 TE:1 loading buffer solution] on an agarose gel along with markers (uncut lambda 1xbet best casino website at concentrations of 10, 20, 50, and 80 ng). Stain gel with 0.5 μg/ml of ethidium bromide for 15 minutes and photograph. Visually compare the intensity of the 1xbet best casino website bands with the markers to determine the concentration.