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

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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