User:Alexander S Mikheyev/Notebook/rotifer alien genes/2010/04/26

Alien genes

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AMPure Cleaned PCR Product 07April10 Ethanol Cleaned Product (~30ul) - Consumed 10ul of remaining Product - Used Standard Agencourt AMPureXP Protocol http://www.beckmangenomics.com/products/dna_purification_and_cleanup/agencourt_ampure_xp.html Final Elution step, only could retrieve ~30ul cleanly, minimal bead pick up. Sample ID Nucleic Acid Conc. Unit A260 A280 260/280 260/230 Sample Type lco/hco 1 0.5 ng/µl 0.01 0.005 1.91 3.78 DNA lco/hco 9 1 ng/µl 0.02 0.016 1.26 2.17 DNA lco/hco 17 -0.1 ng/µl -0.003 -0.017 0.17 0.15 DNA lco/hco 25 1.5 ng/µl 0.03 0.008 3.98 0.84 DNA lco/hco 33 1.2 ng/µl 0.025 0.02 1.25 1.73 DNA lco/hco 41 0.7 ng/µl 0.014 -0.014 -1 1.66 DNA lco/hco 49 1.7 ng/µl 0.035 0.022 1.56 1.21 DNA lco/hco 57 1.2 ng/µl 0.024 0.013 1.82 1.07 DNA lco/hco 65 1.5 ng/µl 0.03 0.028 1.08 1.25 DNA lco/hco 73 1.7 ng/µl 0.034 0.014 2.5 1.19 DNA ' 10ul Ethanol Cleaned -> Total PCR Product Recovery AMPure Cleaned = 30ul lco/hco 1 97 15 lco/hco 9 340 30 lco/hco 17 223 10 lco/hco 25 299 45 lco/hco 33 287 36 lco/hco 41 564 21 lco/hco 49 597 51 lco/hco 57 370 36 lco/hco 65 283 45 lco/hco 73 431 51 PEG Cleaned PCR Product Clean remaining 07April10 Ethanol Cleaned PCR Product ~18ul with PEG. To make PEG: 3g PEG 8000 2.19g NaCl Bring up to 15ml with good water. Lasts 30 days Protocol: 1. Add 1:1 volume of PEG to PCR product. 2. Vortex briefly. 3. Warm at 37degrees for 15minutes. 4. Microcentrifuge for 15minutes at 4000rpm (cold is best). 5. Pipette off supernatant. 6. Wash pellet with 100ul 80% cold ethanol. 7. Spin for 10minutes at 4000rpm. Cold is best. 8. Pipette off ethanol. 9. Repeat steps 6-8 once or twice. 10. Dry down. 11. Resuspend in 10ul good water. 12. NanoDrop 1 0.2 ng/µl 0.004 -0.011 -0.38 0.17 DNA 50 9 0.4 ng/µl 0.007 -0.013 -0.53 0.71 DNA 50 17 0.5 ng/µl 0.01 -0.016 -0.6 0.3 DNA 50 25 2.8 ng/µl 0.057 0.023 2.5 0.37 DNA 50 33 0.4 ng/µl 0.008 -0.011 -0.71 0.89 DNA 50 41 1.5 ng/µl 0.03 0 -1118.82 0.44 DNA 50 49 4.1 ng/µl 0.082 0.035 2.32 0.96 DNA 50 57 2.7 ng/µl 0.054 0.027 2 0.98 DNA 50 65 0.9 ng/µl 0.018 0.001 25.44 0.54 DNA 50 73 1.9 ng/µl 0.039 0.005 7.24 0.87 DNA 50 76 1.8 ng/µl 0.035 -0.003 -10.14 0.61 DNA 50