Brandon:Protocols/Lysis buffers: Difference between revisions
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>Bsos m (→lysis buffer 1) |
>Bsos m (→lysis buffers) |
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Line 8: | Line 8: | ||
100mM NaCl | 100mM NaCl | ||
30mM MgCl2 | 30mM MgCl2 | ||
1% NP40 | 1% IGEPAL® CA-630 (replaces NP40) | ||
*make 1M solution of NaCl and MgCl2 | *make 1M solution of NaCl and MgCl2 | ||
1M | 1M NaCl, 1.461 grams in nuclease free/DI water and bring to 25 mL | ||
1M MgCl2, | 1M MgCl2, 2.3805 grams in nuclease free/DI water and bring to 25 mL (add mgcl2 to water, very exothermic!!) | ||
*To make 10 mL of lysis buffer | *To make 10 mL of lysis buffer | ||
Line 18: | Line 18: | ||
1000 uL of 1M NaCl | 1000 uL of 1M NaCl | ||
333 uL of 1M MgCl2 | 333 uL of 1M MgCl2 | ||
100 uL of IGEPAL® CA-630 (1% in soln, replaces NP40) | |||
8.067 mL Nuclease free water | 8.067 mL Nuclease free water | ||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''''' | |||
| align="center" style="background:#f0f0f0;"|'''''' | |||
| align="center" style="background:#f0f0f0;"|'''10X''' | |||
| align="center" style="background:#f0f0f0;"|'''100X''' | |||
| align="center" style="background:#f0f0f0;"|'''50X''' | |||
| align="center" style="background:#f0f0f0;"|'''40X''' | |||
| align="center" style="background:#f0f0f0;"|'''''' | |||
|- | |||
| Tris-HCl pH 7.5||2.0M||500||5000||2500||2000||uL | |||
|- | |||
| NaCl||5M||200||2000||1000||800||uL | |||
|- | |||
| MgCl||1M||333||3330||1665||1332||uL | |||
|- | |||
| IGEPAL CA-630||solid||100||1000||500||400||uL | |||
|- | |||
| ||||||0||0||0||uL | |||
|- | |||
| |||||||||||| | |||
|- | |||
| |||||||||||| | |||
|- | |||
| ||||||11330||5665||4532||uL | |||
|- | |||
| ||||H20||-1330||4335||5468||uL | |||
|- | |||
| | |||
|} |
Latest revision as of 20:03, 21 November 2014
lysis buffers[edit]
lysis buffer 1[edit]
- Crawford et al. PNAS 2003
- lysis buffer composition:
10X lysis buffer 100mM Tris.Hcl pH 7.5 100mM NaCl 30mM MgCl2 1% IGEPAL® CA-630 (replaces NP40)
- make 1M solution of NaCl and MgCl2
1M NaCl, 1.461 grams in nuclease free/DI water and bring to 25 mL 1M MgCl2, 2.3805 grams in nuclease free/DI water and bring to 25 mL (add mgcl2 to water, very exothermic!!)
- To make 10 mL of lysis buffer
500 uL of 2M Tris.Hcl PH7.5 1000 uL of 1M NaCl 333 uL of 1M MgCl2 100 uL of IGEPAL® CA-630 (1% in soln, replaces NP40) 8.067 mL Nuclease free water
' | ' | 10X | 100X | 50X | 40X | ' |
Tris-HCl pH 7.5 | 2.0M | 500 | 5000 | 2500 | 2000 | uL |
NaCl | 5M | 200 | 2000 | 1000 | 800 | uL |
MgCl | 1M | 333 | 3330 | 1665 | 1332 | uL |
IGEPAL CA-630 | solid | 100 | 1000 | 500 | 400 | uL |
0 | 0 | 0 | uL | |||
11330 | 5665 | 4532 | uL | |||
H20 | -1330 | 4335 | 5468 | uL | ||