Protocol Online: Cached

1x; pH7.4; 7.6; 8.0; (store at 4^oC).

	Conc.	Stock	1ml	5ml	10ml	50ml	400ml
Tris Cl	10mM	1M	10µl	50µl	100µl	500µl	4ml
EDTA	1mM	0.5M	2µl	10µl	20µl	100µl	800µl
H₂O		mQ	988µl	4.94ml	9.88ml	49.4ml	395ml

10x:

	Conc.	Stock	1ml	5ml
Tris Cl	0.1M	1M	0.1ml	0.5ml
EDTA	10mM	0.5M	20µl	0.1ml
H₂O	mQ		880µl	4.4ml

autoclave sterilization

STE

1x; (store at 4^oC).

	Conc.	Stock	1ml	5ml	10ml	50ml
NaCl	0.1M	5M	20µl	100µl	200µl	1.0ml
Tris Cl, pH8.0	10mM	1M	10µl	50µl	100µl	0.5ml
EDTA, pH8.0	1mM	0.5M	2µl	10µl	20µl	100µl
H₂O		mQ	0.968ml	4.84ml	9.68ml	48.4ml

10x:

	Conc.	Stock	1ml	5ml
NaCl	1.0M	5M	0.2ml	1.0ml
Tris Cl	0.1M	1M	0.1ml	0.5ml
EDTA	10mM	0.5mM	20µl	0.1ml
H₂O	mQ		680µl	3.4ml

autoclave sterilization

SSC

20x; pH 7.0; (store at NT).

	Conc.	Stock	0.5L	1.5L	2L
NaCl	3M	58.44g/M	87.66g	263.0g	350.6g
Na₃C₆H₅O₇x 5.5H₂O	0.3M	357.1g/M	53.57g	160.7g	214.3g
H₂O		mQ	440.3ml	1.32L	1.76L

p = 1.103

TAE

1x pH = 7.6:

Tris-acetate 40mM
EDTA 1mM

store the large stock at 4^oC, working quantity (~100ml) at NT

	Conc.	Stock	100ml	150ml	200ml	250ml
Tris-base	2M	121.1g/M	24.22g	36.33g	48.44g	60.55g
EDTA	0.05M	372.3g/M	1.862g	2.792g	3.723g	4.654g
Acetic acid	~1.56M	17.4M	~8.96ml ~9.40g	~13.44ml ~14.10g	~17.92ml ~18.80g	~22.40ml ~23.50g	~ ~
H₂O		mQ	~73.30g	~109.95g	~146.60g	~183.25g	~

p(50x)=1.0878

it is convenient to solubilize Tris and EDTA first, and only then add acid to adjust pH.

TBE

10x, pH ~8.3;(store at NT in clean bottle)

	1x	10x	Stock	1L
Tris base	89mM	0.89M	121.14g/M	107.8g
Boric acid	89mM	0.89M	61.83g/M	55.03g
EDTA, pH 8.0	2mM	20mM	500mM	40ml 43.84g
H₂O			mQ	863.3ml

10x stock solution is stable at NT if to clean it through 0.4µm filter;

for PAAG it is used as 1x, for agarose gel - as 0.5x;

for agarose gels it is used for separation of small (< 200bp) fragments. Generally accepted, that the purification of fragments is more complex from TBE gels.

FGRB

50x (formaldehyde gel-running buffer), p=1.104g/ml (store at +4^oC):

	Conc.	Stock	0.5L	1.0L	2.0L
MOPS (pH7.0)	1.0M	209.27g/M	104.64g	209.27g	418.54g
AcONax3H₂O	0.5M	136.08g/M	34.02g	68.04g	136.08g
EDTAx2H₂O	50mM	372.3g/M	9.31g	18.62g	37.23g
H₂O		mQ	404.0ml	808ml	1616ml

	Conc.	Stock	0.5L	1.0L	2.0L
MOPS (pH7.0)	1.0M	209.27g/M	104.64g	209.27g	418.54g
AcONa anhydrous	0.5M	82.04g/M	20.51g	41.02g	82.04g
EDTA x 2H₂O	50mM	372.3g/M	9.31g	18.62g	37.23g
H₂O		mQ	417.5ml	835ml	1670ml

adjust pH7.0 10N NaOH (per 500ml of buffer ~22ml), filter-sterilize through 0.22µm filter;

solution becomes yellow during the storage on the light, but it has no effect on the solution quality.

GTGB

20x /Glycerol tolerant gel buffer/ (store at +4^oC):

	Conc.	Stock	1L	1.5L	2.0L	2.5L
Tris base	1.78M	121.14g/M	216g	324g	432g	540g
Taurine	0.56M	125.15g/M	72g	108g	144g	180g
Na₂EDTAx2H₂O	10.7mM	372.3 g/M	4g	6g	8g	10g
H₂O		mQ	802ml	1203ml	1604ml	2005ml

p=1.094

Taurine: C₂H₇NO₃S; Mw=125.15g/M.

TGB

Tris-Glycine buffer 5x, pH8.3, (store the main stock at +4^oC, working quantity at NT).

	1x	Conc.	Stock	1L	1.5L
Tris-base	25mM	125mM	121.1g/M	15.1g	22.71g
Glycine	250mM	1.25M	75.05g/M	94g	141g
SDS	0.1%	0.5%	10%	50ml	75ml
H₂O			mQ

Loading buffer

10x, (store at NT).

	Conc.	Stock	1ml	5ml	10ml	25ml
Xilene cyanol	0.1%	solid	1.0mg	5.0mg	10mg	25.0mg
Bromph. blue	0.1%	solid	1.0mg	5.0mg	10mg	25.0mg
SDS	0.5%	10%	50µl	250µl	500µl	1.25ml
EDTA, pH8.0	0.1M	0.5M	0.2ml	1.0ml	2.0ml	5.0ml
Glycerol	50%	100%	0.5ml 0.625g	2.5ml 3.125g	5.0ml 6.25g	12.5ml 15.63g
H₂O		mQ	0.25ml	1.25ml	2.5ml	6.25ml

PEG-NaCl

(store at 4^oC).

	Conc.	Stock	50ml	100ml	150ml	200ml
NaCl	1.6M	58.44g/M	4.675g	9.35g	14.03g	18.70g
PEG-6000	13%	solid	6.50g	13.00g	19.50g	26.00g

or:

	Conc.	Stock	50ml	100ml	150ml	200ml
NaCl	1.6M	5M	16ml 18.97g	32ml 37.95g	48ml 56.92g	64ml 75.90g
PEG-6000	13%	40%	16.25ml	32.50ml	48.75ml	65.00ml

(store at 4^oC).

	Conc.	Stock	10ml	50ml	100ml	150ml
NaCl	0.1M	5M	200µl	1.0ml	2.0ml	3.0ml
MgSO₄*	10mM	1M	100µl	0.5ml	1.0ml	1.5ml
Tris Cl, pH7.5	50mM	1M	0.5ml	2.5ml	5.0ml	7.5ml
	0.01%	2%	50µl	0.25ml	0.5ml	0.75ml
H₂O		mQ	9.15ml	45.75ml	91.5ml	137.25ml

* - add after autoclaving;

autoclave sterilization

STM

(store at 4^oC):

	Conc.	Stock	50ml
NaCl	10mM	5M	100µl
Tris Cl pH8.0	50mM	1M	2.5ml
MgCl₂	10mM	1M	500µl
H₂O		mQ	46.9ml

(store at 4^oC):

	Conc.	Stock	50ml
Tris Cl pH8.0	50mM	1M	2.5ml
MgCl₂	10mM	1M	500µl
H₂O		mQ	47.0ml

Denaturation buffer

(store at NT).

	Conc.	Stock	100ml	150ml	300ml	500ml	750ml
NaCl	1.5M	58.44g/M	8.766g	13.15g	26.30g	43.83g	65.75g
NaOH	0.5M	10M	5ml 6.65g	7.5ml 9.97g	15ml 19.94g	25ml 33.23g	37.5ml 49.84g
		40g/M	2g	3g	6g	10g	15g

Neutralization buffer

(store at NT).

	Conc.	Stock	150ml	300ml	500ml	750ml
Tris Cl, pH7.4	1M	121.1g/M	18.15g	36.3g	60.5g	90.75g
NaCl	1.5M	58.44g/M	13.15g	26.30g	43.83g	65.75g
HCl	~0.81M	11.6M/L	~10.5ml	~21ml	~35ml	~52.5ml
EDTA (optional)	1mM	0.5M	0.3ml	0.6ml	1ml	1.5ml

Developer Rentgen-2

(store at NT in the dark).

	500ml	800ml	1l
	1.1g	1.76g	2.2g
Na₂SO₃ anhidr.	36.0g	57.6g	72.0g
Hydroquinone	4.4g	7.04g	8.8g
Na₂CO₃	24.0g	38.4g	48.0g
KBr	2.0g	3.2g	4.0g
H₂O	490ml	784ml	980ml

solubilize in the presented order , p=1.115g/ml.

Fixer BKF-2

(store at NT).

	500ml	800ml	1l
Na₂S₂O₃x 5H₂O	130g	208g	260g
NH₄Cl	25g	40g	50g
Sodium pyrosulfite	8.5g	13.6g	17g
H₂O	398ml	636.8ml	796ml

solubilize in the presented order, p=1.123g/ml.

Scintillation solution

(store at NT in the dark):

	0.5L	1L
PPO	2.50g	5.00g
POPOP	0.05g	0.10g
Toluene

Plasmid solution I

(store at +4^oC)

	Conc.	Stock	50ml	100ml	150ml	200ml
Glucose	50mM	2M	1.25ml 1.41g	2.5ml 2.825g	3.75ml 4.24g	5.0ml 5.56g
Tris Cl, pH 8.0	25mM	1.0 M	1.25ml	2.5ml	3.75ml	5.0ml
EDTA	10mM	0.5 M	1.0ml	2.0ml	3.0ml	4.0ml
H₂O		mQ	46.5ml	93ml	139.5ml	186ml

Plasmid solution II

	Conc.	Stock	0.5ml	1ml	2.5ml	5.0ml	6.5ml	8.5ml
NaOH	0.2N	1N	0.1ml	0.2ml	0.5ml	1.0ml	1.3ml	1.7ml
SDS	1%	10%	0.05ml	0.1ml	0.25ml	0.5ml	0.65ml	0.85ml
H₂O		mQ	0.350ml	0.7ml	1.75ml	3.5ml	4.55ml	5.95ml

Oligonucleotide gel staining mix

(store at NT).

	Conc.	Stock	100ml	200ml	300ml	500ml
AcONa(5.2)	0.5M	3M	17ml	33ml	50.0ml	83ml