Editing Protein Expression By Autoinduction (section)

== ZYM-505 medium for plasmid preparation ==

{| border="1" cellspacing="1" cellpadding="1" style="width: 500px;"
|-
| <br/>
| <br/>
| <br/>
| Final
|-
| 968 ml<br/>
| ZY<br/>
| 
tryptone

yeast extract

| 
1%&nbsp;

0.5%

|-
| 20 ml<br/>
| &nbsp;50 x M<br/>
| 
Na2HPO4&nbsp;

KH2PO4&nbsp;

(NH4)2SO4&nbsp;

Na2SO4

| 
25 mM&nbsp;

25 mM

50 mM

5 mM

|-
| 10 ml<br/>
| &nbsp;50 x 505<br/>
| 
glycerol&nbsp;

glucose&nbsp;

| 
0.5%&nbsp;

0.05%&nbsp;

|-
| 2 ml<br/>
| &nbsp;1 M MgSO4<br/>
| MgSO4<br/>
| 2 mM<br/>
|-
| 0.2 ml<br/>
| 1000 x trace elements<br/>
| trace elements<br/>
| 0.2X<br/><br/>
|}

=== '''ZY''' ===

1% tryptone: 10 g/L

0.5% yeast extract:&nbsp; 5 g/L



=== '''50 x M''' ===

1.25 M Na2HPO4-7H2O 335 g/L

1.25 M KH2PO4 170 g/L

2.5 M NH4Cl 134 g/L

0.25 M Na2SO4 35.5 g/L

Add in sequence to 700 ml water and stir until all salts have been dissolved.Occasionally crystals appear but they can be re-dissolved in the microwave. ThepH of a 20-fold dilution in water should be ~6.75.

=== '''100 x 5055''' ===

glycerol 500 g/L

glucose 50 g/L

=== '''20 x P''' ===

1.0 M Na2HPO4-7H2O 268 g/L

1.0 M KH2PO4 136 g/L

0.5 M (NH4)2SO4 66 g/L

Add in sequence to 770 ml water and stir until all salts have been dissolved. The pH of a 20-fold dilution in water should be ~6.75.

=== '''50 x 5052''' ===

25% glycerol 250 g/L

2.5% glucose 25 g/L

10% alpha-lactose 100 g/L&nbsp; α-lactose

Add in sequence to 730 ml water and stir until all components have been dissolved. Lactose is slow to dissolve. It may take two hours or more at room temperature. The process can be sped up by heating in microwave.

=== '''1000x trace elements''' ===

50 mM FeCl3

20 mM CaCl2

10 mM MnCl2

10 mM ZnSO4

2 mM CoCl2

2 mM CuCl2

2 mM NiCl2

2 mM Na2MoO4

2 mM Na2SeO3

2 mM H3BO3

Preparation of 100 ml 1000x trace elements stock solution (in ~60 mM HCl).

Prepare stock solutions of the salts (except FeCl3) as mentioned in the table below.

Dissolve 0.1 M FeCl3 in 50 ml of 100x diluted concentrated HCl.

Add to 36 ml sterile water: 50 ml 0.1 M FeCl3-6H2O 2.70 g/100 ml

2 ml 1.0 M CaCl2-2H2O 15.8 g/100 ml

1 ml 1.0 M MnCl2-4H2O 19.8 g/100 ml

1 ml 1.0 M ZnSO4-7H2O 28.8 g/100 ml

1 ml 0.2 M CoCl2-6H2O 4.76 g/100 ml

2 ml 0.1 M CuCl2-2H2O 1.70 g/100 ml

1 ml 0.2 M NiCl2-6H2O 4.76 g/100 ml

2 ml 0.1 M Na2MoO4-2H2O 2.42 g/100 ml

2 ml 0.1 M Na2SeO3 1.73 g/100 ml

2 ml 0.1 M H3BO3 0.62 g/100 ml

{| border="1" cellspacing="1" cellpadding="1" style="width: 500px;"
|-
| <br/>
| <br/>
| <br/>
| Final
|-
| 928 ml<br/>
| ZY<br/>
| 
tryptone

yeast extract

| 
1%&nbsp;

0.5%

|-
| 50 ml<br/>
| &nbsp;20 x P<br/>
| 
Na2HPO4&nbsp;

KH2PO4&nbsp;

(NH4)2SO4&nbsp;

| 
50 mM&nbsp;

50 mM

25 mM

|-
| 20 ml<br/>
| &nbsp;50 x 5052<br/>
| 
&nbsp;glycerol&nbsp;

glucose&nbsp;

alpha-lactose&nbsp;

| 
0.5%&nbsp;

0.05%&nbsp;

0.2%

|-
| 2 ml<br/>
| &nbsp;1 M MgSO4<br/>
| MgSO4<br/>
| 2 mM<br/>
|-
| 0.2 ml<br/>
| 1000 x trace elements<br/>
| trace elements<br/>
| 0.2X<br/><br/>
|}
[[Category:Protocols]]