- Product Overview
- Catalog
- Workflows
- prepGEM
- > Original
- > Bacteria
- > Blood
- > Storage Card Blood
- > Fungi
- > Insect
- > Retrieval
- > Saliva
- > Storage Card Saliva
- > Tissue
- forensicGEM
- livestockGEM
- Applications
- Publications & Posters
DNA Extraction
Using
prepGEM™ Tissue
prepGEM™ Tissue is a kit specifically formulated for extracting quality DNA from a range of animal tissue types including: muscle, fat, rat tails, and fish fins. The extracted DNA is suitable for PCR applications including qPCR, SNP detection and routine genotyping.
- The kit uses optimized methods and formulations to provide DNA in less than 20 minutes.
- Reactions are closed-tube and hands-off.
- All buffers and reagents are PCR compatible, so no downstream purification is required.
These features make prepGEM™ Tissue ideal for automated sample processing with most liquid handling platforms for full or partial automation. The few steps and simple methodology of prepGEM™ Tissue also allow for manual processing of hundreds of samples, or single sample preps.
To purchase products over the phone or by fax, proceed to customer service >>
Workflow
Extraction Method
The following procedure outlines general guidelines. Different sample types will require slight method variations.
Preparation
All manipulations should be performed in a clean room or a PCR hood. Use only certified DNA-free tubes and reagents and wash forceps, scalpels and dissection surfaces in 0.05% hypochlorite bleach. Rinse thoroughly with DNA-free water.
|
Tissue Type |
Procedure
|
| Animal tissue (fat, muscle, organs) | Cut the tissue into cubes of approximately 1.5 mm3. Mash the sample with the side of the scalpel and place in a thin-walled PCR tube. |
| Hair follicles | Cut the shaft 5 mm above the follicle and place in a thin-walled PCR tube. |
| Rat tail | Cut the shaft 5 mm above the follicle and place in a thin-walled PCR tube. |
| Fish fin/tail | Use approximately 4 mm2 of a fish tail or fin. Place in a thin-walled PCR tube. |
1. Add:
- 89 µl DNA-free water
- 10 µl of 10x buffer GOLD
- 1 µl prepGEM™
2. Incubate at 75°C for 15 minutes.
3. Incubate at 95°C for 5 minutes. A thermal cycler can be used for this step
4. Aspirate the extract away from the tissue.
Technical Tips
- The prepGEM™ Tissue procedure can be automated using most liquid handling robots.
- prepGEM™ Tissue is a preparative method for DNA extraction - it is not a purification protocol. Its purpose is to lyse cells and to strip the DNA of nucleoproteins. The formulation creates DNA that can be used for SNiPs, STRs, quantitative, multiplex and routine PCR applications.
- For accurate yield assessment, a qPCR is recommended. The DNA produced by prepGEM™ is approximately 90% single-stranded. If standard fluorescent chelating dyes are to be used for quantification, then this factor should be taken into consideration.
- As with any preparative method for nucleic acid extraction, for best results prepare and manage samples at 4ºC, or on ice, before and after extraction.
- When storing the sample after extraction, aspirate the supernatant from the precipitated residue and store separately.
- prepGEM™ is stable for 30 days at 4º C, for longer-term storage prepGEM™ should be stored at -20ºC.
- For long-term storage of the extracted DNA, add TE buffer to 1x and store at -20°C.
Typical Results
| MT16SF1 | ACCCCGCCTGTTTACCAA |
| MT16SR4 | CTGATCCAACATCGAGGTCG |
| COWF1 | CTGGCAAAGTGGACATCGTC |
| COWF2 | TCCATTGATGACGAGCTTCC |
Figure 1: qPCR plots from beef fat nuclear and mitochondrial primers.
L14735 |
AAA AAC CAC CGT TGT TAT TCA ACT A |
H15149 |
GCI CCT CAR AAT GAY ATT TGT CCT CA |
Figure 2: Frozen filleted fish-fingers (Hoki) using mitochondrial primers for species identification.