N-BANDING PROTOCOL
The following protocol describes the standard N-banding protocol routinely used in our laboratory for chromosome identification in 1xbet best casino website and 1xbet best casino website relatives and will give satisfactory results even in the hands of an inexperiencesd worker.
- Germinate seeds in petri dishes on moist filter paper for 3–4 days.
- Collect root tips when 1.5–2.5-cm long and pretreat for 24 h in an ice-1xbet best casino website bath.
- Fix in1xbet bestat room temperature for 1–3 days and then freeze or refrigerate until use.
- Stain root tips in 1% acetocarmine for 1–2 h at room temperature, or overnight in at 4°C.
- Make squash preparations in 45% acetic acid.
- Remove cover slips by either the dry ice method, dipping 1xbet best casino website into liquid nitrogen for about 10 sec, or placing 1xbet best casino website into a freezer (-70°C) for 3 min.
- Incubate 1xbet best casino website in hot 45% acetic acid for 3 min in a water bath at 50°C.
- Transfer 1xbet best casino website to 95% ethanol at room temperature for 10 min.
- Air dry 1xbet best casino website overnight.
- Incubate 1xbet best casino website in phosphate buffer (120 g NaH2PO4/ L distilled 1xbet best casino website ) for 2 min at 94°C in a 1xbet best casino website bath.
- Rinse in tap 1xbet best casino website and air dry.
- Stain with Leishman's stain (BDH #R03087) in Soerensen's phosphate buffer for about 20 min. The staining solution is diluted 5 X with 1xbet best casino website and Leishman's stain added to a final concentration of 15%).
Soerensen's buffer Stock Stain Part A sodium phosphate dibasic (Na2HOP4) 9.47 g / L H2O 58 mL Part B potassium dihydrogen phosphate (KH2PO4) 9.07 g / L H2O
- Rinse briefly in tap 1xbet best casino website and air dry.
- For recording of N-bands, a cover slip can be mounted using a drop of xylene or preparations can be observed without a cover slip using the appropriate lenses.
