Luckau Protocols:Tris-Cl: Difference between revisions
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[[Luckau_Protocols | Tara's Protocols]] | [[Luckau_Protocols | Tara's Protocols]] | ||
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==Purpose== | ==Purpose== | ||
*Tris-Cl is a buffering agent (made from Tris base and HCl) | *Tris-Cl is a buffering agent (made from Tris base and HCl) | ||
* | *In the Clark Lab, 0.5M Tris-Cl, pH 8.0 is used as a stock solution for Low TE | ||
*In the Clark Lab, 1M Tris-Cl, pH 8.0 is used as a stock solution for Low TE | |||
*In the Clark Lab, 1M Tris-Cl, pH 8.3 is used as a stock solution for making the PCR Buffer series (A through H) | |||
==Info== | ==Info== | ||
*tris [hydroxymethyl] amino methane (C<sub>4</sub>H<sub>11</sub>NO<sub>3</sub>, 2-Amino-2-hydroxymethyl-propane-1,3-diol) | *tris [hydroxymethyl] amino methane (C<sub>4</sub>H<sub>11</sub>NO<sub>3</sub>, 2-Amino-2-hydroxymethyl-propane-1,3-diol) | ||
*MW = 157.56 g/mol | :: we currently use [http://www.amresco-inc.com/TRIS-0497.cmsx Amresco, product code 0497], Lot 2901C127 | ||
*MW = 157.56 g/mol | |||
* Note: when adjusting pH, be patient: adjust the pH slowly until it seems steady and stable, then leave the solution on the stirrer for an hour or so, then re-check the pH to be sure it hasn't changed. It's always a good idea to re-re-check the pH a day after as well. | |||
==To make <math>\tfrac{1}{2}</math> liter of 0.5M Tris-Cl, pH 8.0== | |||
===Calculations=== | ===Calculations=== | ||
* <math>500 \mathrm{mL} \bullet \tfrac{0.5 \mathrm{mol}}{\mathrm{L}}\ \bullet \tfrac{\mathrm{L}}{1000 \mathrm{mL}}\ \bullet \tfrac{157.56 \mathrm{g}}{\mathrm{mol}}\ = 39.39 \mathrm{g}</math> | |||
===Protocol=== | |||
# Using digital scale, place 39.39 g Tris-base powder into weight boat | |||
# In 1 L beaker, mix Tris-base powder in 400 mL H<sub>2</sub>O | |||
# Put beaker on hotplate with stirbar until dissolved | |||
# Use concentrated aqueous HCl to adjust to pH 8.0 (remember to be patient: adjust, let it sit, adjust again) | |||
# Add H<sub>2</sub>O to bring volume to 500 mL | |||
# Re-check pH | |||
# Autoclave solution, store at room temperature | |||
==To make <math>\tfrac{1}{2}</math> liter of 1M Tris-Cl, pH 8.0== | |||
=== | ===Calculations=== | ||
* <math>500 \mathrm{mL} \bullet \tfrac{1 \mathrm{mol}}{\mathrm{L}}\ \bullet \tfrac{\mathrm{L}}{1000 \mathrm{mL}}\ \bullet \tfrac{157.56 \mathrm{g}}{\mathrm{mol}}\ = 78.78 \mathrm{g}</math> | |||
# In 1 L beaker, mix Tris- | ===Protocol=== | ||
# Using digital scale, place 78.78 g Tris-base powder into weight boat | |||
# In 1 L beaker, mix Tris-base powder in 400 mL H<sub>2</sub>O | |||
# Put beaker on hotplate with stirbar until dissolved | # Put beaker on hotplate with stirbar until dissolved | ||
# | # Use concentrated aqueous HCl to adjust to pH 8.0 (remember to be patient: adjust, let it sit, adjust again) | ||
# Add H<sub>2</sub>O to bring volume to 500 mL | # Add H<sub>2</sub>O to bring volume to 500 mL | ||
# Re-check pH | |||
# Autoclave solution, store at room temperature | |||
==To make 250 mL of 1M Tris-Cl, pH 8.3== | |||
===Calculations=== | |||
* <math>250 \mathrm{mL} \bullet \tfrac{1 \mathrm{mol}}{\mathrm{L}}\ \bullet \tfrac{\mathrm{L}}{1000 \mathrm{mL}}\ \bullet \tfrac{121.14 \mathrm{g}}{\mathrm{mol}}\ = 30.285 \mathrm{g}</math> | |||
===Protocol=== | |||
# Using digital scale, place 30.285 g Tris-base powder into weight boat | |||
# In 250 mL glass media bottle, mix Tris-base powder in approximately 200 mL H<sub>2</sub>O | |||
# Put beaker on stirrer with stirbar until dissolved | |||
# Use concentrated aqueous HCl to adjust to pH 8.3 (remember to be patient: adjust, let it sit, adjust again) | |||
# Add H<sub>2</sub>O to bring volume to 250 mL | |||
# Re-check pH; the next day, re-check it again | |||
# Autoclave solution, store at room temperature | # Autoclave solution, store at room temperature |
Latest revision as of 15:44, 27 September 2012
Tris-Cl Protocol |
Purpose
- Tris-Cl is a buffering agent (made from Tris base and HCl)
- In the Clark Lab, 0.5M Tris-Cl, pH 8.0 is used as a stock solution for Low TE
- In the Clark Lab, 1M Tris-Cl, pH 8.0 is used as a stock solution for Low TE
- In the Clark Lab, 1M Tris-Cl, pH 8.3 is used as a stock solution for making the PCR Buffer series (A through H)
Info
- tris [hydroxymethyl] amino methane (C4H11NO3, 2-Amino-2-hydroxymethyl-propane-1,3-diol)
- we currently use Amresco, product code 0497, Lot 2901C127
- MW = 157.56 g/mol
- Note: when adjusting pH, be patient: adjust the pH slowly until it seems steady and stable, then leave the solution on the stirrer for an hour or so, then re-check the pH to be sure it hasn't changed. It's always a good idea to re-re-check the pH a day after as well.
To make [math]\displaystyle{ \tfrac{1}{2} }[/math] liter of 0.5M Tris-Cl, pH 8.0
Calculations
- [math]\displaystyle{ 500 \mathrm{mL} \bullet \tfrac{0.5 \mathrm{mol}}{\mathrm{L}}\ \bullet \tfrac{\mathrm{L}}{1000 \mathrm{mL}}\ \bullet \tfrac{157.56 \mathrm{g}}{\mathrm{mol}}\ = 39.39 \mathrm{g} }[/math]
Protocol
- Using digital scale, place 39.39 g Tris-base powder into weight boat
- In 1 L beaker, mix Tris-base powder in 400 mL H2O
- Put beaker on hotplate with stirbar until dissolved
- Use concentrated aqueous HCl to adjust to pH 8.0 (remember to be patient: adjust, let it sit, adjust again)
- Add H2O to bring volume to 500 mL
- Re-check pH
- Autoclave solution, store at room temperature
To make [math]\displaystyle{ \tfrac{1}{2} }[/math] liter of 1M Tris-Cl, pH 8.0
Calculations
- [math]\displaystyle{ 500 \mathrm{mL} \bullet \tfrac{1 \mathrm{mol}}{\mathrm{L}}\ \bullet \tfrac{\mathrm{L}}{1000 \mathrm{mL}}\ \bullet \tfrac{157.56 \mathrm{g}}{\mathrm{mol}}\ = 78.78 \mathrm{g} }[/math]
Protocol
- Using digital scale, place 78.78 g Tris-base powder into weight boat
- In 1 L beaker, mix Tris-base powder in 400 mL H2O
- Put beaker on hotplate with stirbar until dissolved
- Use concentrated aqueous HCl to adjust to pH 8.0 (remember to be patient: adjust, let it sit, adjust again)
- Add H2O to bring volume to 500 mL
- Re-check pH
- Autoclave solution, store at room temperature
To make 250 mL of 1M Tris-Cl, pH 8.3
Calculations
- [math]\displaystyle{ 250 \mathrm{mL} \bullet \tfrac{1 \mathrm{mol}}{\mathrm{L}}\ \bullet \tfrac{\mathrm{L}}{1000 \mathrm{mL}}\ \bullet \tfrac{121.14 \mathrm{g}}{\mathrm{mol}}\ = 30.285 \mathrm{g} }[/math]
Protocol
- Using digital scale, place 30.285 g Tris-base powder into weight boat
- In 250 mL glass media bottle, mix Tris-base powder in approximately 200 mL H2O
- Put beaker on stirrer with stirbar until dissolved
- Use concentrated aqueous HCl to adjust to pH 8.3 (remember to be patient: adjust, let it sit, adjust again)
- Add H2O to bring volume to 250 mL
- Re-check pH; the next day, re-check it again
- Autoclave solution, store at room temperature