FavorPrep 96-well Gel/PCR Clean-Up Kit User Manual
Contents
1. Please Read Important Notes Before Starting The Following Steps Step1 Transfer up to 100 mg of agarose gel containing relevant DNA fragment to each well of a clean 96 Well 2 ml Plate not provided Add 300 ul of Binding Buffer D1 to each well sealing with a adhesive film Incubate at 50 C for 10 15 minutes until the gel slice dissolved completely During the incubation briefly shake the incubated plate for every 5 minutes to make the sample mixture mix well with Binding Buffer D1 Step 2 DNA Binding Place a 96 Well DNA Binding Plate on top of the vacuum manifold Transfer the sample mixture from step 1 to each well of the 96 Well DNA Binding Plate Apply vacuum at 10 inches Hg for 3 minutes until wells have emptied vacuum at 10 inches Hg 3 min Step 3 Washing Add 650 ul of Wash Buffer ethanol added to each well of the 96 Well DNA Binding Plate Apply vacuum at 10 inches Hg for 5 minutes until wells have emptied Apply vacuum at 10 inches Hg for an additional 10 min or incubate at 60 C for 10 min to remove residual ethanol incubate at 65 C for 5 min vacuum at 10 inches Hg or 15 min STEP 4 DNA Elution Place a clean 96 well PCR Plate provided on top of a clean 96 Well 2 ml Plate not provided And place the 96 Well DNA Binding Plate on top of the clean 96 Well PCR plate top 96 well DNA Binding Plate middle 96 well PCR Plate bottom 96 Well 2 ml plate Add 50 75 ul of Elu2. mg agarose gel slice Binding Capacity up to 20 ug well DNA Size range 70 bp 12Kb Operation centrifuge vacuum manifold Handling Time about 30 minutes for PCR clean up about 40 minutes for gel DNA extraction Recovery 90 95 PCR clean up 70 85 for gel DNA extraction Downstream application Fluorescent or radioactive sequencing Restriction digestion Library screening Ligation Labeling Transformation Important Note 1 Buffer provided in this kit contain irritants Wear gloves and lab coat when handling these buffers 2 When excising the agarose gel remove the extra gel to minimize the size of the gel 3 Preheat a water bath to 50 C for Gel DNA Extraction Protocol 4 Add 200 ml of ethanol 96 100 to each Wash Buffer before first open 1 Gel DNA Extraction Protocol Centrifuge Please Read Important Notes Before Starting The Following Steps Step1 Transfer up to 100 mg of agarose gel containing relevant DNA fragment to each well of a clean 96 Well 2 ml Plate not provided Add 300 ul of Binding Buffer D1 to each well sealing with a adhesive film Incubate at 50 C for 10 15 minutes until the gel slice dissolved completely During the incubation briefly shake the incubated plate for every 5 minutes to make the sample mixture mix well with Binding Buffer D1 Step 2 DNA Binding Place a 96 Well DNA Binding Plate on top of a 96 Well 2 ml Plate not provided Transfer the sample mixture from step 1 to the 96 W
3. FavorPrep 96 well Gel PCR Clean Up Kit User Manual Cat No FAPKE 001 FAPKE 002 For Research Use Only v 1211 1 Introduction FavorPrep 96 well GEL PCR Clean Up kit is designed for rapid purification of fragment DNA from agarose gel PCR or other enzymatic reaction The procedure uses chaotropic salt to dissolve agarose and denature enzyme With the suitable binding condition provided by this system the DNA in the sample mixture binds to glass fiber matrix in the 96 well DNA Binding Plate The contaminants are washed with an ethanol contained wash buffer and finally the purified DNA is eluted by low salt Elution buffer or water The protocol does not require phenol extraction and alcohol precipitation The entire procedure can be done within 30 40 minutes and the purified DNA is ready for restriction digestion ligation labeling PCR and sequencing reaction Quality Control The quality of 96 Well Gel PCR Kit is tested on a lot to lot basis The purified DNA fragment is checked by agarose gel analysis and quantified with spectrophotometer Kit Content FAPKEOO1 FAPKEOO2 Bind Buffer D1 130 ml 330 ml Wash Buffer concentrated 50 ml 50 mi x 3 Elution Buffer 30 ml 60 ml 96 Well DNA Binding plate 4 pcs 10 pcs 96 Well PCR plate 4 pcs 10 pcs Adhesive Film 5 pcs 12 pcs Add 200 ml of ethanol 96 100 to each Wash Buffer when first use Specification Sample Size up to 50 ul PCR or other enzymatic reaction mixture Up to 100
4. ell DNA Binding Plate Place the assembly plates in a rotor bucket and centrifuge at 4 500 6 000 x g for 5 min Discard the flow through and return the 96 Well DNA Binding Plate to the 96 Well 2 ml Plate 4 500 6 000 x g 5 min STEP 3 Washing Add 650 ul of Wash Buffer ethanol added to each well of the 96 Well DNA Binding Plate Place the assembly plates in a rotor bucket and centrifuge at 4 500 6 000 x g for 5 min Discard the flow through and return the 96 Well DNA Binding Plate back to the 96 Well 2 ml Plate Place the assembly plates in a rotor bucket and centrifuge at 4 500 6 000 xg for an additional 15 minutes to remove residual ethanol remove residual ethanol 4 500 6 000 x g 4 500 6 000 x g 5 min 15 min STEP 4 DNA Elution Place a clean 96 well PCR Plate provided on top of a clean 96 Well 2 ml Plate not provided And place the 96 Well DNA Binding Plate on top of the clean 96 Well PCR plate top 96 well DNA Binding Plate middle 96 well PCR Plate bottom 96 Well 2 ml plate Add 50 75 ul of Elution Buffer or ddH20 pH 8 0 8 5 on the membrane center of the 96 Well DNA Binding Plate Stand for 3 minutes until Elution Buffer or ddH20 has been absorbed by the membrane completely Place the combined plates in a rotor bucket and centrifuge for 5 min at 4 500 6 000 x g for 5 min to elute purified DNA 4 500 6 000 x g 5 min Gel DNA Extraction Protocol Vacuum
5. inches Hg for an additional 10 min or incubate at 60 C for 10 min to remove residual ethanol incubate at 65 C for 5 min vacuum at 10 inches Hg or 15 min STEP 4 DNA Elution Place a clean 96 well PCR Plate provided on top of a clean 96 Well 2 ml Plate not provided And place the 96 Well DNA Binding Plate on top of the clean 96 Well PCR plate top 96 well DNA Binding Plate middle 96 well PCR Plate bottom 96 Well 2 ml plate Add 50 75 ul of Elution Buffer or ddH20 pH 8 0 8 5 on the membrane center of the 96 Well DNA Binding Plate Stand for 3 minutes until Elution Buffer or ddH20 has been absorbed by the membrane completely Place the combined plates in a rotor bucket and centrifuge for 5 min at 4 500 6 000 x g for 5 min to elute purified DNA
6. ol 4 500 6 000xg 7 5 min STEP 4 DNA Elution Place a clean 96 well PCR Plate provided on top of a clean 96 Well 2 ml Plate not provided And place the 96 Well DNA Binding Plate on top of the clean 96 Well PCR plate top 96 well DNA Binding Plate middle 96 well PCR Plate bottom 96 Well 2 ml plate Add 50 75 ul of Elution Buffer or ddH2O pH8 0 8 5 on the membrane center of the 96 Well DNA Binding Plate Stand for 3 minutes until Elution Buffer or ddH2O0 has been absorbed by the membrane completely Place the combined plates in a rotor bucket and centrifuge for 5 min at 4 500 6 000 x g for 5 min to elute purified DNA 4 500 6 000 x g 5 min PCR Clean Up Protocol Vaccum Please Read Important Notes Before Starting The Following Steps Step1 Transfer up to 50 pl of PCR or enzymatic product to each well of a clean 96 Well 2 ml Plate not provided Add 250 ul of Binding Buffer D1 to each well Mix well by Pipetting Step 2 DNA Binding Place a 96 Well DNA Binding Plate on top of the vacuum manifold Transfer the sample mixture from step 1 to each well of the 96 Well DNA Binding Plate Apply vacuum at 10 inches Hg for 3 minutes until wells have emptied vacuum at 10 inches Hg 3 min Step 3 Washing Add 650 ul of Wash Buffer ethanol added to each well of the 96 Well DNA Binding Plate Apply vacuum at 10 inches Hg for 5 minutes until wells have emptied Apply vacuum at 10
7. tion Buffer or ddH20 pH8 0 8 5 on the membrane center of the 96 Well DNA Binding Plate Stand for 3 minutes until Elution Buffer or ddH2O has been absorbed by the membrane completely Place the combined plates in a rotor bucket and centrifuge for 5 min at 4 500 6 000 x g for 5 min to elute purified DNA 4 500 6 000 x g 5 min PCR Clean Up Protocol Centrifuge Please Read Important Notes Before Starting The Following Steps Step1 Transfer up to 50 pl of PCR or enzymatic product to each well of a clean 96 Well 2 ml Plate not provided Add 250 ul of Binding Buffer D1 to each well Mix well by Pipetting Step 2 DNA Binding Place a 96 Well DNA Binding Plate on top of a 96 Well 2 ml Plate not provided Transfer the sample mixture from step 1 to the 96 Well DNA Binding Plate Place the assembly plates in a rotor bucket and centrifuge at 4 500 6 000 x g for 5 min Discard the flow through and return the 96 Well DNA Binding Plate to the 96 Well 2 ml Plate STEP 3 Washing Add 650 ul of Wash Buffer ethanol added into each well of the 96 Well DNA Binding Plate Place the assembly plates in a rotor bucket and centrifuge at 4 500 6 000 x g for 5 min Discard the flow through and return the 96 Well DNA Binding Plate back to the 96 Well 2 ml Plate Place the assembly plates in a rotor bucket and centrifuge at 4 500 6 000 xg for an additional 15 minutes to remove residual ethanol remove residual ethan
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