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        Recipe for yeast tryptone medium

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        Recipe for yeast tryptone medium

        Keywords: Zappe; YT 2x; Yeast tryptone; E. coli
        Recipe for yeast tryptone medium 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
        Recipe for yeast tryptone medium 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
        Recipe for yeast tryptone medium 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
        Recipe for yeast tryptone medium 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
        Recipe for yeast tryptone medium 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
        Recipe for yeast tryptone medium 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
        Recipe for yeast tryptone medium Recipes: BAL-31 dilution/storage buffer Contributor: Zappe, H Related: Bal31 shortening Storage: -20 C Bal31 dilution and storage buffer Component stock conc final conc. / 10ml Tris-HCl , pH 8.0 1M 20mM 0.2ml CaCl2 1M 5mM 50 ml MgCl2 1M 5mM 50 ml EDTA , pH 8.0 0.5M 1mM 20 ml NaCl 5M 0.1M 0.20ml Water - 5.58ml glycerol 44%(v/v) 4.40ml
        
        

        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
        Recipe for yeast tryptone medium 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
        Recipe for yeast tryptone medium 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.



        Recipe for TE Buffer

        Keywords: TE buffer; Babb
        Recipe for yeast tryptone medium 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
        Recipe for yeast tryptone medium 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
        Recipe for yeast tryptone medium Chloroform:isoamyl alcohol mix

        Contributor: Zappe, H

        Related: many protocols

        Care:

        Recipe for yeast tryptone medium Recipe for yeast tryptone medium caution

        Chloroform:Isoamyl (24:1)

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



        Recipe for cysteine

        Keywords: cysteine
        Recipe for yeast tryptone medium 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
        Recipe for yeast tryptone medium 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
        



        DNA digestion: restriction enzyme core reaction buffer

        Keywords: Zappe; DNA; digestion; restriction
        Recipe for yeast tryptone medium 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
        Recipe for yeast tryptone medium 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
        Recipe for yeast tryptone medium 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.



        Recipe for DNA extraction lysing solution

        Keywords: Lysing solution; DNA extraction; Zappe
        Recipe for yeast tryptone medium Lysing solution

        Contributor: Zappe, H

        Related: Bacterial DNA extraction

      • Lysing solution (2X):

        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.



        Recipe for DTT

        Keywords: Zappe; DTT; dithiothreitol
        Recipe for yeast tryptone medium 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



        Recipe for EDTA

        Keywords: EDTA; ethyl???; Zappe
        Recipe for yeast tryptone medium 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



        Recipe for ethidium bromide

        Keywords: Zappe; EtBr; ethidium bromide
        Recipe for yeast tryptone medium Ethidium bromide (EtBr)

        Contributor: Zappe, H

        Related: many protocols

        Care:

        Recipe for yeast tryptone medium never breath dust - mutagen

        Ethidium bromide (EtBr)

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





        Recipe for ethanol stock

        Keywords: ethanol; EtOH
        Recipe for yeast tryptone medium Ethanol (EtOH)

        Contributor: Zappe, H

        Related: many protocols

        Ethanol (EtOH)

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