Quick Guide Rev. 4 - 22/12/16
ION-Liquid Force Extraction Buffer
Content:
Type:
EXD008
10 ml buffered solution*
Qualitative
Shipping:
< 40°C
Storage:
2-8°C
Expiry date:
[1]
See date on the packaging, product validity refers to the product kept
intact in its original packaging and constantly under suitable temperature
conditions as mentioned above.
* reagents are supplied with an 5% of extra volume.
Bacterial DNA extraction from enriched samples
For the extraction of DNA from bacterial pellet derived from the centrifugation of enrichment broth.
1. Collect 1.5 ml of the enriched complex matrix in a 2.0 ml (or 1.5 ml) conical tubes and centrifuge
at 2.000 rpm for 2 min in order to collect only coarse particulates on the bottom of the tube (do
not exceeding with centrifuge speed and time).
This step may be skipped if enriched media results fairly clean: thus, proceed with step 2 as follow.
2. Collect 1 ml of supernatant (or withdraw 1 ml of enriched media if skipped step 1) using a pipette
and transfer to a new 1,5 ml conical tube (supernatant may be still opaque due to cells
presence).
3. Centrifuge at 14.000 rpm for 5 min to pellet cells.
4. Discard the supernatant and accurately remove any residual broth over the cell pellet. Leave a
small volume remaining (~100 μl) if an obvious pellet is not observed.
5. Add 100 µl of GENERlex ION-Liquid Force Extraction Buffer.
6. Vortex the pellet to easy its resuspension.
7. Incubate the tube for 10 min at 95 ± 3°C.
8. Add DNA-free water to achieve 1 ml of DNA elute volume: in the sample you will find DNA
corresponding to initial enriched sample volume and in this way you will avoid PCR inhibition.
9. DNA can be use immediately for qPCR reaction or stored frozen until use.
If the Internal Amplification Control (IAC) of the Real-Time PCR kit in use shows no-amplification or
amplification plots grossly abnormal (PCR inhibition) we advise to test samples diluted 1:10.
Use DNase/RNase Free Water to prepare dilutions.
Warning and Precaution
Do not mix components between different bottles even when having same lot numbers. This assay is
designed to be used by laboratory personnel following the usual molecular biology precautions i.e. GLP.
Disclaimers
Generon S.p.A. guarantees the buyer exclusively concerning the quality of reagents and of the
components used to manufacture the product. Generon S.p.A. is not responsible and cannot anyway be
considered responsible or jointly responsible for any possible damages resulting from the utilization of the
product by the user and from the data obtained.
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