Synthesis PHP-CBA-DSP-PEI18
Synthesis of PHP
[edit]1.5 g solid 1,3-Diamino-2-propanol, MW 90.12, 16.6 mmole
+3 ml MeOH
+3.63 g 1,3-dibrono-2-propanol , MW 217.89, 16.7 mmole
50 oC for 48 hours
store at -20 oc.
add 5 ml H2O to dissolve
+ 25 ml EtOH, mix, spin 3000 rpm for 5 min
remove supernatant
Add 5 ml H2O, + 25 ml EtOH
spin, repeat 2 more times
yield: 2.8 g wet PHP + H2O to 10 ml
Synthesis PHP-DSP-PEI
[edit]3 ml PHP solution, + 25 mg PEI18, + 25 mg DSP, 40 oC reaction o/n
9-26-2011
4.5 ml PHP solution, + 37 mg PEI18, + 37 mg DSP, 40 oC reaction o/n
Synthesis PHP-CBA-PEI
[edit]3 ml PHP solution + 25 mg CBA, + 25 mg PEI18, 40 oC reaction o/n
Guanydinylation
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Transfection
[edit]Make 1:10, 1:30, 1:40, 1:50, 1:75, 1:100 dilution
use 3 ul diluted polymer sol, + 1 ug DNA, 50 ul DMEM for 12 well plate. incubation time: 20 min
1: PHP-CBA-PEI, 1:10
2: PHP-CBA-PEI, 1:30
3: PHP-CBA-PEI, 1:40
4: PHP-CBA-PEI, 1:50
5: PHP-CBA-PEI, 1:75
6: PHP-CBA-PEI, 1:100
7: PHP-DSP-PEI, 1:10
8: PHP-DSP-PEI, 1:30
9: PHP-DSP-PEI, 1:40
10:PHP-DSP-PEI, 1:50
11: PHP-DSP-PEI, 1:75
12: PHP-DSP-PEI, 1:100
Results
[edit]12 hour after transfection
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Day 1 after transfection
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Day 2 after transfection
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Day 3 after transfection
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July 17, 2011