Preparing Solutions

Short Form

Obtain the required amount of ingredients

Dissolve them Bring to volume (q.s.) Store

Short Form

Obtain the required amount of ingredients

Dissolve them Bring to volume (q.s.) Store

Getting started

Beaker larger than final volume

Add 2/3 final volume of gdH2O

Add a magnetic spin bar

Begin adding reagents

Use a clean spatula and weigh dish for each ingredient

Never return excess material to its container

Issues

Heating pH Solvents Noxious and

hazardous compounds

Short Form

Obtain the required amount of ingredients

Dissolve themBring to volume (q.s.) Store

“Finishing” a solution

Everything should be fully dissolved*

Temperature must be cool enough to handle.

pH must be set Transfer to graduated

cylinder and bring to final volume

Final volume = q.s. (quantum satis)

Late Edition

(Should be late addition) Filter sterilized amendments Heat sensitive, reactive (e.g. ampicillin) 1000X – Volume insignificant

Short Form

Obtain the required amount of ingredients

Dissolve them Bring to volume (q.s.)Store

Issues – “Begin with the end in mind”

Autoclaving Filtering Light Heat Containers

Labels

Composition - 20X SSC (better exact composition)

(special) Storage conditions Date Made (include the year!) Your name Autoclave tape

Short Form

Obtain the required amount of ingredients

Dissolve them Bring to volume (q.s.) Store

Powders – Molarity; Three numbers

(g mol-1) MW, FW (hydrates), % purity, free acid vs. salt, etc.

(mol l-1)(g mol-1)(l) = grams required

Powders – % (w/v)

Grams of powder added per 100 ml final volume (q.s. rules)

1% = g 100 ml-1

Liquids – Molarity; Density

Divide g required by density of liquid (g ml-1)

Pipette this amount of liquid Remember to account for purity.

(mol l-1)(g mol-1)(l) = grams required

Additions from concentrated stock solutions

C1 is the concentration of your stock solution

C2 is the concentration you want in the end

V2 is your final volume (q.s.) V1 is how much to add!

C1V1 = C2V2

Additions from concentrated stock solutions

Make sure concentration units are the same (M, mM, %, etc.)

Make sure volumes are in the same units (l, ml, etc.)

Solve for V1

C2 cannot be bigger than C1

V1 cannot be bigger than V2

C1V1 = C2V2

X solutions

20X SSC Buffer 5X Wash Solution 10X RE Buffer 20X TAE Buffer 100X Vitamins 1000x Ampicillin

X refers to relative concentration of some complex solution

1X is the normal working concentration Use C1V1 = C2V2 to determine amount needed (V1)

of concentrated stock (C1)

A Note on Buffers

Buffer refers to the salt of a weak acid or base that helps to keep a constant pH.

Buffer is used to describe solutions containing pH buffers.

E.g. 10X PCR buffer contains 10 times of everything required for the PCR reaction including Tris, a pH buffer

Adding the Buffer

Tris, Acetate, Phosphate Add buffer to give desired

concentration Add acid (or base) to adjust pH to

desired value Bring to volume

0.5 l of 50 mM Tris-HCl, pH 7.9

(0.05 mol l-1)(121.1 g mol-1)(0.5 l) = 3.03g Add 3.03 g Tris (base) to about 400

ml of gdH2O Add 6 M HCl dropwise to adjust pH

to 7.9 Bring to 500 ml in graduated

cylinder

Multi Component Solutions...20% (wt/vol) sucrose-0.3 M Tris-HCl (pH 8)-1mM EDTA....

Start with ca. 600 ml water Add _____g sucrose, spin to

completely dissolve Add _____g Tris base, spin to

completely dissolve Add _____ml of 250 mM Stock EDTA

pH 8 solution Adjust pH to 8.0 with _____. Bring to 1000 ml total volume.