کیت استخراج DNA مایع آمنیوتیک|DNA Isolation From Amniotic Fluid
Amniotic Fluid DNA Isolation Kit
This kit should be stored at room temperature.
If properly stored, all kit components are stable until the expiration date printed on the label.
After adding absolute ethanol to AM5 Buffer, Store the AM5 Buffer at +2 to +8°C
Additional Equipment and Reagent required
- Absolute ethanol
- Standard tabletop microcentrifuge capable of 13,000 x g centrifugal force
- Microcentrifuge tubes, 1.5 ml, sterile
This kit provides a convenient and rapid method to isolate total DNA from fresh, frozen and preserved Amniotic Fluid Samples. The purified DNA is of the highest quality and is fully compatible with all downstream applications such as PCR, qPCR, NGS and microarrays since all humic acid substances and other PCR inhibitors are removed during the isolation process.
The procedure is optimized to achieve reliable results within 12 min.
Handling Requirements and Safety Information
All solutions are clear and should not be used when precipitates have formed. Warm the solutions at +15 to +25°C or in a 37°C water bath until the precipitates have dissolved.
AM4 Buffer contain guanidinium hydrochloride which is an irritant.
Do not allow AM4 Buffer to touch your skin, eyes, or mucous membranes. If contact does occur, wash the affected area immediately with large amounts of water.
If you spill the reagent, dilute the spill with water before wiping it up.
Do not use any modified ethanol.
Do not pool reagents from different lot numbers.
Immediately after usage, close all bottles in order to avoid leakage, varying buffer concentrations or buffer conditions.
After first opening store all bottles in an upright position.
Do not allow the AM4 Buffer to mix with sodium hypochlorite found in commercial bleach solutions. This mixture can produce a highly toxic gas.
Wear protective disposable gloves, laboratory coats and eye protection, when handling samples and kit reagents.
Do not contaminate the reagents with bacteria, virus, or nucleases. Use disposable pipets and nuclease free pipet tips only, to remove aliquots from reagent bottles.
Working Solution Preparation
Add 4 ml absolute ethanol to AM5 Buffer Bottle.
Add 8 ml absolute ethanol to AM5 Buffer Bottle.
Add 16 ml absolute ethanol to AM5 Buffer Bottle.
Please note that the Ethanol concentration of a AM5 Buffer may decrease during long term storage resulting in a drop-down of the final DNA yield.
before starting Incubate the EB buffer at 55 to 65 ° C until the end of the protocol to obtain the maximum yields.
- Add 1 ml AM1 Buffer to the 2ml microtube and add 800 μl Amniotic Fluid sample. Mix gently (don’t Vortex) and centrifuge at maximum speed for 1 minutes.
- Discard the supernatant and add 1ml AM1 Buffer again. Mix gently (don’t Vortex) and centrifuge at maximum speed for 1 minutes.
- Discard the supernatant and Add 300 μl AM2 Buffer. Pipette completely.
- Add 300 μl Am3 Buffer then add 600 μl Chloroform mix vigorously for 10 seconds (don’t Vortex). Centrifuge at 7000 RPM for 4 min.
- Transfer the supernatant into a 1.5 ml microtube gently. Add 200 μl of AM4 Buffer to the sample. mix gently (don’t Vortex).
- Transfer the Step 5 solution into a DNA spin column assembled in a clean collection tube (provided). Centrifuge at maximum speed for 1 minutes.
- Disconnect the DNA spin column from collection tube and discard the flow through solution. Reconnect the DNA spin column to the collection tube.
- Add 200 μl AM5 Buffer to DNA spin column and centrifuge at maximum speed for 1 minutes. Discard flow through.
- Place the DNA spin column into the clean new microtube. Add 50 μl of pre-warmed EB Buffer or sterile DDW directly onto column membrane and stand for 3 min. centrifuge at maximum speed for 1 minutes.
- Check 7-10 μl of extracted DNA by 1% agarose gel electrophoresis. Store DNA at -20 ̊C