Recipe for yeast tryptone medium

Recipe for yeast tryptone medium

Keywords: Zappe; YT 2x; Yeast tryptone; E. coli

Yeast tryptone medium

Contributor: Zappe, H

Related: E. coli growth

Yeast tryptone broth/agar, 2xYT

Component  stock conc final conc. /1l
Tryptone - - 16g
Yeast extract - - 10g
NaCl - - 5g

for agar plates

agar - - 15g

autoclave.

Recipe for ammonium acetate

Keywords: ammonium acetate; Zappe

Ammonium acetate

Contributor: Zappe, H

Related: many protocols

Storage: 4 C

Ammonium acetate

Component  stock conc final conc. / 200 ml /1l
Ammonium acetate - 7.5M 289g 
water - - to 200ml

Can be used as Solution III for plasmid preparation.

Recipe for Ammonium persulphate

Keywords: Zappe; APS; ammonium persulphate

Ammonium persulphate

Contributor: Zappe, H

Related: SDS-PAGE gels

Storage: Make fresh weekly

Ammonium persulphate (APS)

Component  stock conc final conc. / 0.5ml
APS  10%(w/v) 0.05g
water   0.5ml

Recipe fo ampicillin

Keywords: ampicillin; Zappe

Ampicillin

Contributor: Zappe, H

Related: many protocols

Storage: -20 C

Ampiciliin (Sigma, sodium salt), 1000x

Component  stock conc final conc. / 20 ml /1l
Ampicillin - 100 mg/ml 2g 
water - - to 20ml

filter sterilize and store aliquots at -20 C

dilute 1/1000 for final conc of 100 mg/ml

Recipe for ATP

Keywords: ATP; Zappe

ATP

Contributor: Zappe, H

Related: many protocols

Storage: -20 C

ATP (0.1M)

Component  stock conc final conc. / 0.8ml ml /1l
ATP - 0.1M 60mg

NaOH

 0.1M - adjust to pH 7.0
water - - to 0.8ml

Adjust to pH 7.0 with 0.1M NaOH (spotting a few ml on pH paper)

Freeze 0.1ml aliquots

Recipe for Bal31 nuclease reaction buffer

Keywords: Zappe; Bal31; DNA shortening; buffer

Recipes: BAL-31 reaction buffer (5X).

Contributor: Zappe, H

Related: Bal31 shortening

Storage: 4 C

Bal31 reaction buffer, 5x

Component  stock conc final conc. / 10ml

Tris-HCl

, pH 8.0 1M 0.1M 1ml

CaCl2

 1M 60mM 0.6ml

MgCl2

 1M 60mM 0.6ml

EDTA

, pH 8.0 0.5M 5mM 0.1ml

NaCl

 5M 3M 6ml
Water  - 1.7ml

Recipe for Bal31 nuclease dilution/storage buffer

Keywords: Zappe; Bal31; DNA shortening; buffer

Recipes: BAL-31 dilution/storage buffer Contributor: Zappe, H Related:

All the components, except the glycerol, are combined in a 10 ml sterilin tube. The tube is then filled to the 10 ml mark with sterile glycerol. Mix by gently shaking the tube. The volume marks on the sterilin tube are used as it is not possible to accurately pipette 5 ml of undiluted glycerol.

Recipe for TA buffer

Keywords: TA buffer; ligation; Zappe; electrophoresis

Recipes: TA buffer

Contributor: Zappe, H

Related: Gel based DNA manipulation

TA buffer

Component  stock conc final conc. / 10ml

Tris-HCl

, pH 8.0 1M 

NaAcl

 5M 
Water  ???- 5.58ml

??

Recipe for Tris-borate buffer

Keywords: Zappe; TB; electrophoresis; gel

TB buffer

Contributor: Zappe, H

Related: Gel electrophoresis

Tris-borate buffer (5x)

Component  stock conc final conc. / 1000ml
Tris-base   54g
Boric acid   27.5g

EDTA

, pH 8.0 0.5M  20ml

Do not store Tris-borate in a glass bottle.

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Recipe for TE Buffer

Keywords: TE buffer; Babb

TE buffer

Contributor: Zappe, H

Related: many protocols

TE Buffer

Component  stock conc final conc. / 10 ml /100ml /1l

Tris

, pH 8.0 1M 10mM 0.1ml 1ml
 10ml

EDTA

, pH 8.0 0.5M 1mM 20ml 0.2ml
 2ml ??

Recipe for calcium chloride

Keywords: Zappe; calcium chloride; cacl2

Calcium chloride

Contributor: Zappe, H

Related: many protocols

Storage: 4 C

Calcium chloride (1M), CaCl2

Component  stock conc final conc. / 100 ml /1l
CaCl2.2H20 - 1M 14.7g 
water - - to 100ml

autoclave

Recipe for chloroform:isoamyl alcohol mixture

Keywords: chloroform:isoamyl alcohol mix; Babb

Chloroform:isoamyl alcohol mix

Contributor: Zappe, H

Related: many protocols

Care:

caution

Chloroform:Isoamyl (24:1)

Component  stock conc final conc. / 10 ml /100ml
Chloroform - - - 96ml
Isoamyl alcohol - - - 4ml

Recipe for cysteine

Keywords: cysteine

Recipes: Cysteine

Contributor: Zappe, H

Related: -

Cysteine

Component  stock conc final conc. / 10 ml
Cysteine 0.1M 1mM 0.1ml
Water  - to 4.69ml

DNA digestion: restriction enzyme dilution buffer

Keywords: Zappe; DNA; digestion; restriction

Restriction enzyme dilution buffer

Contributor: Zappe, H

Related: DNA digestion

Storage: -20 C

Restriction enzyme dilution buffer

Component  stock conc final conc. / 10ml

Tris-HCl

, pH 7.5 1M 10mM 0.1ml

NaCl

 5M 50mM 0.1ml
b mercapto-ethanol  10mM 7 ml
Gelatin 10mg/ml 100 mg/ml 0.1ml
water   5.3ml
glycerol  44%(v/v) 4.4 ml

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DNA digestion: restriction enzyme core reaction buffer

Keywords: Zappe; DNA; digestion; restriction

Restriction enzyme core buffers (10X).

Contributor: Zappe, H

Related: DNA digestion

Storage: -20 C

Restriction enzyme dilution buffer (10x)

Salt concentration mM

Component  stock conc 0 50 100 150

Tris-HCl

, pH 7.5 1M 1ml 1ml 1ml
 1ml

NaCl

 5M 0 1ml 2ml
 87.7mg
BSA (albumin) 10mg/ml 1ml 1ml 1ml 1ml

MgCl2

 1M 1ml 1ml 1ml 1ml

DTT

 0.5M 0.2ml 0.2ml 0.2ml 0.2ml
water  2.4ml 1.4ml 0.4ml 2.4ml
glycerol  44%(v/v) 4.4 ml 4.4 ml 4.4ml

DNA digestion: Sma I restriction enzyme reaction buffer

Keywords: Zappe; DNA; digestion; restriction; Sma I

Sma I restriction enzyme reaction buffer

Contributor: Zappe, H

Related: DNA digestion

Storage: -20 C

Sma I restriction enzyme reaction buffer (10x)

Component  stock conc final conc. / 10ml

Tris-HCl

, pH 8.0 1M 0.1M 1ml

KCl

 5M 0.2M 2ml

MgCl2

 1M 0.2M 2ml

DTT

 0.5M 0.2ml 0.2ml
water   0.4ml
glycerol  44%(v/v) 4.4ml

All the components, except the glycerol, are combined in a 10 ml sterilin tube. The tube is then filled to the 10 ml mark with sterile glycerol. Mix by gently shaking the tube. The volume marks on the sterilin tube are used as it is not possible to accurately pipette 5 ml of undiluted glycerol.

Recipe for DNA extraction cell suspension buffer

Keywords: Cell suspension buffer; DNA extraction; Zappe

Cell suspension buffer

Contributor: Zappe, H

Related: Bacterial DNA extraction

Cell suspension buffer:

Component  stock conc final conc. / 100 ml /1l /10ml

Tris-HCl

 (pH 8.0) 1M 10mM 1ml

EDTA

 (pH 8.0) 0.5M 1mM 0.2ml
sucrose - 0.35M  12g
water - - to 100ml

If the cells are going to be physically disrupted (French press or glass beads), then do not add the sucrose.

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Recipe for DNA extraction lysing solution

Keywords: Lysing solution; DNA extraction; Zappe

Lysing solution

Contributor: Zappe, H

Related: Bacterial DNA extraction

Component  stock conc final conc. / 100 ml /1l /10ml

Tris-HCl

 (pH 8.0) 1M 100mM 10ml 

EDTA

 (pH 8.0) 0.5M 20mM 4ml

NaCl

 5M 0.3M 6ml

SDS

 25%(w/v) 2% 8ml
b-mercaptoethanol - 2%(v/v) 2ml
proteinase K 20mg/ml 100mg/ml 0.5ml
water - - to 100ml

Add proteinase K (50 *l of a 20 mg/ml stock solution per 10 ml 2X lysing solution) and the *- mercaptoethanol to the amount of lysing solution needed. The other Components can be premixed as a stock solution.

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Recipe for DTT

Keywords: Zappe; DTT; dithiothreitol

Dithiothreitol (DTT)

Contributor: Zappe, H

Related: many protocols

Storage: -20 C

Dithiothreitol (DTT)

Component  stock conc final conc. / 20 ml /1l
DTT - 1M 3.09g 

sodium acetate

, pH 5.2 - - to 
20ml

filter sterilize and store aliquots at -20 C

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Recipe for EDTA

Keywords: EDTA; ethyl???; Zappe

EDTA (ethyl??)

Contributor: Zappe, H

Related: many protocols

EDTA (Ethylene??)

Component  stock conc final conc. / 400 ml /1l
EDTA - 0.5M 93.05g 
NaOH - - +/- 10g
water - - to 400ml

EDTA will not dissolve until +/- pH 8.0

autoclave

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Recipe for ethidium bromide

Keywords: Zappe; EtBr; ethidium bromide

Ethidium bromide (EtBr)

Contributor: Zappe, H

Related: many protocols

Care:

never breath dust - mutagen

Ethidium bromide (EtBr)

Component  stock conc final conc. / 10 ml /1l
EtBr - 10mg/ml 0.1g 
water - - to 10ml

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Recipe for ethanol stock

Keywords: ethanol; EtOH

Ethanol (EtOH)

Contributor: Zappe, H

Related: many protocols

Ethanol (EtOH)

Component  stock conc final conc. / 500 ml /1l
EtOH - 70% 350ml 
water - - 150ml