Arichard:Reagents/celseq primers: Difference between revisions
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>Andrew (Created page with "==CEL-Seq RT primer layout== ===Stock layout=== * Barcoded RT primers are in 48 wells of a 92 well plate at 200 uM each, in the following layout: {| {{table}} | align="cent...") |
>Andrew No edit summary |
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===Diluted primer layout=== | ===Diluted primer layout=== | ||
# Dilute primers to 32 pmol/ul, 3.2 pmol/ul, and 0.32 pmol/ul, depending on input (Note that each primer must be diluted separately for each sample): | |||
## Add 16 ul of 100 pmol/ul primer to 84 ul H2O --> 100 ul 32 pmol/ul primer. Use 0.5 ul of this per 1 ng total RNA. | |||
## Add 10 ul of 32 pmol/ul primer to 90 ul H2O --> 100 ul 3.2 pmol/ul primer. Use 0.5 ul of this per 100 pg total RNA. | |||
## Add 10 ul of 3.2 pmol/ul primer to 90 ul H2O --> 100 ul 0.32 pmol/ul primer. Use 0.5 ul of this per 10 pg total RNA. | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''100 ul/well''' | |||
| align="center" style="background:#f0f0f0;"|'''1''' | |||
| align="center" style="background:#f0f0f0;"|'''2''' | |||
| align="center" style="background:#f0f0f0;"|'''3''' | |||
| align="center" style="background:#f0f0f0;"|'''4''' | |||
| align="center" style="background:#f0f0f0;"|'''5''' | |||
| align="center" style="background:#f0f0f0;"|'''6''' | |||
| align="center" style="background:#f0f0f0;"|'''7''' | |||
| align="center" style="background:#f0f0f0;"|'''8''' | |||
| align="center" style="background:#f0f0f0;"|'''9''' | |||
| align="center" style="background:#f0f0f0;"|'''10''' | |||
| align="center" style="background:#f0f0f0;"|'''11''' | |||
| align="center" style="background:#f0f0f0;"|'''12''' | |||
|- | |||
| Conc||32 pmol/ul||3.2 pmol/ul||0.32 pmol/ul||32 pmol/ul||3.2 pmol/ul||0.32 pmol/ul||32 pmol/ul||3.2 pmol/ul||0.32 pmol/ul||32 pmol/ul||3.2 pmol/ul||0.32 pmol/ul | |||
|- | |||
| A||#1||#1||#1||#2||#2||#2||#3||#3||#3||#4||#4||#4 | |||
|- | |||
| B||#5||#5||#5||#6||#6||#6||#7||#7||#7||#8||#8||#8 | |||
|- | |||
| C||#9||#9||#9||#10||#10||#10||#11||#11||#11||#12||#12||#12 | |||
|- | |||
| D||#13||#13||#13||#14||#14||#14||#15||#15||#15||#16||#16||#16 | |||
|- | |||
| E||#17||#17||#17||#18||#18||#18||#19||#19||#19||#20||#20||#20 | |||
|- | |||
| F||#21||#21||#21||#22||#22||#22||#23||#23||#23||#24||#24||#24 | |||
|- | |||
| G||#25||#25||#25||#26||#26||#26||#27||#27||#27||#28||#28||#28 | |||
|- | |||
| H||#29||#29||#29||#30||#30||#30||#31||#31||#31||#32||#32||#32 | |||
|- | |||
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Latest revision as of 22:11, 14 February 2013
CEL-Seq RT primer layout[edit]
Stock layout[edit]
- Barcoded RT primers are in 48 wells of a 92 well plate at 200 uM each, in the following layout:
Barcode number | 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | 10 | 11 | 12 |
A | 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | 10 | 11 | 12 |
B | 13 | 14 | 15 | 16 | 17 | 18 | 19 | 20 | 21 | 22 | 23 | 24 |
C | 25 | 26 | 27 | 28 | 29 | 30 | 31 | 32 | 33 | 34 | 35 | 36 |
D | 37 | 38 | 39 | 40 | 41 | 42 | 43 | 44 | 45 | 46 | 47 | 48 |
E | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty |
F | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty |
G | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty |
H | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty | Empty |
- Serial dilutions are necessary to dilute the RT primers to the correct concentration.
Diluted primer layout[edit]
- Dilute primers to 32 pmol/ul, 3.2 pmol/ul, and 0.32 pmol/ul, depending on input (Note that each primer must be diluted separately for each sample):
- Add 16 ul of 100 pmol/ul primer to 84 ul H2O --> 100 ul 32 pmol/ul primer. Use 0.5 ul of this per 1 ng total RNA.
- Add 10 ul of 32 pmol/ul primer to 90 ul H2O --> 100 ul 3.2 pmol/ul primer. Use 0.5 ul of this per 100 pg total RNA.
- Add 10 ul of 3.2 pmol/ul primer to 90 ul H2O --> 100 ul 0.32 pmol/ul primer. Use 0.5 ul of this per 10 pg total RNA.
100 ul/well | 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | 10 | 11 | 12 |
Conc | 32 pmol/ul | 3.2 pmol/ul | 0.32 pmol/ul | 32 pmol/ul | 3.2 pmol/ul | 0.32 pmol/ul | 32 pmol/ul | 3.2 pmol/ul | 0.32 pmol/ul | 32 pmol/ul | 3.2 pmol/ul | 0.32 pmol/ul |
A | #1 | #1 | #1 | #2 | #2 | #2 | #3 | #3 | #3 | #4 | #4 | #4 |
B | #5 | #5 | #5 | #6 | #6 | #6 | #7 | #7 | #7 | #8 | #8 | #8 |
C | #9 | #9 | #9 | #10 | #10 | #10 | #11 | #11 | #11 | #12 | #12 | #12 |
D | #13 | #13 | #13 | #14 | #14 | #14 | #15 | #15 | #15 | #16 | #16 | #16 |
E | #17 | #17 | #17 | #18 | #18 | #18 | #19 | #19 | #19 | #20 | #20 | #20 |
F | #21 | #21 | #21 | #22 | #22 | #22 | #23 | #23 | #23 | #24 | #24 | #24 |
G | #25 | #25 | #25 | #26 | #26 | #26 | #27 | #27 | #27 | #28 | #28 | #28 |
H | #29 | #29 | #29 | #30 | #30 | #30 | #31 | #31 | #31 | #32 | #32 | #32 |