BP-10Spin Column Blood Genomic DNA Minipreps Kit

Kit Contents

(a) TBMBuffer may form a precipitate upon storage. If necessary, dissolve theprecipitate by warming at 37^oC.

(b) Before use, add 150ul or 300ul of sterilizedwater to the tube containing 2 mg or 4 mg of proteinase K. Keep at -20^oC. Before use, add 48ml of 100% ethanol to 12 ml Wash Solution for GDMIP-50, or add 96 ml of 100% ethanol to 24 ml Wash Solution for GDMIP-100. Forother volumes of wash solution, simply add enough ethanol to make a 4:1 ratio (volume of added ethanol: volume of Wash Solution = 4:1).

(c) Elution Buffer is 2.0 mMTris-HCl pH 8.0~8.5. Although TE buffer pH 8.0 or water can be used, yield isgenerally 20% lower.

Storage:

With the exception of the Proteinase K, thekit may be stored at room temperature. The proteinase K should be stored at 4ºC. The kit is stable for 12 months at roomtemperature. For longer storage, keepall contents cold.

Principle

This BP-10 spin columnmini-preps kit is designed for a fast isolation of genomic DNA from clinicalsamples such as fresh or frozen whole blood ( with common anticoagulatants suchas citrate, EDTA, and heparin); , serum, bone, and other body fluids, and cellsuspensions. 200ul-400ul per column of healthy whole blood sample is enough toobtain 30ug of pure genomic DNA. Nucleotides, proteins, salts, and other impuritiesdo not bind to the BP-10 Column. Purified blood genomic DNA can be applied inmost molecular biology experiments including restriction digestion, PCR,Southern-blotting etc.

Application:

• Extraction of Genomic DNA from whole blood and other body fluids.

Features:

• High quality of genomic DNA from blood and other clinical samples
• High yields and reproducible
• No phenol / chloroform extraction , no ethanol precipitation

Procedure for Extraction of BloodDNA:

1. Harvest the appropriate 0.5 ml the whole blood in 2.0ml collection tubeby centrifugation at 3000rpm for 3 minutes at 4℃. Discard supernatant.
2. Add 0.8 ml TBP Buffer to the collection tube vortex gently, then 3000rpm for 3 minutes,Discard supernatant.
3. Repeat the step 2. If the blood pellet looks mauve，then continue next step.
4. Add 0.5 ml TBM Buffer to the collection tube, intensity vortex. then add3 ul Proteinase K, Then incubate at 55℃ for 30 minutes.
5. If insoluble material is visible, centrifuge for 2 minutes at 5,000rpm.Save the supernatant to 2.0ml collection tube, then add 260ul absolute Ethanol.
6. Apply the mixture to BP-10 column that is in a 2.0 ml collection tube.Spin at 8,000 rpm for 1 minute.. Discardthe flow-through in the collection tube.
7. Add 500 ml of Wash Solution, and spin at 8,000 rpm for 1 minute.
8. Repeat washing step 7.
9. Discard flow-through. Spin at 8,000rpm for an additional minute toremove residual amount of Wash Solution.
10. Place the column into aclean 1.5 ml microfuge tube. Add 30-50 ul Elution Buffer into the center partof membrane in the column. Incubate the tube at 37 or 50℃ for 2 minutes. Incubate at 37 or 50℃ could increase recoveryyield.
11. Spin at 10,000 rpm for 1minute to elute DNA from the column.

Measure DNA quantity by UV absorption at A₂₆₀ (1.0 OD unit is equivalent of 50 ug). Assessgenomic DNA quality by an analytical 0.7% agarose gel. Isolated genomic DNAshould not contain RNA. Its length should be over 50 kb.