User:Jarle Pahr/Kits

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Current revision (09:44, 21 May 2013) (view source)
 
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||Dh5a:pSB-M1g||21.5.13|| ||~4||LB + Kan 50 ug/mL ||13 mL tube|| ||2.6 |||| || || || ||
||Dh5a:pSB-M1g||21.5.13|| ||~4||LB + Kan 50 ug/mL ||13 mL tube|| ||2.6 |||| || || || ||
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DongSheng Bio Eco friendly plasmid extraction kit:
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Observed plasmid yields:
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{| class="wikitable" border="1" cellpadding="5" cellspacing="0"
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! Strain:Plasmid!! Date !! Culture volume incubated (mL) !! Culture volume centrifuged (mL) !!Medium /Antibiotic !! Incubation vessel  !! Incubation time!! OD600 !! Elution volume (uL) !! Obtained DNA concentration (ng/uL) !! Total yield (ug) !! Yield (ug / mL * OD600) !! 260/280 !! 260/230
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|Strain:Plasmid|| Date ||Culture vol inc||Culture vol centr||medium||incub vessel||incub time||OD600||El. vol.|| conc  || tot || rel || 260/280 || 260/230
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Current revision

Commercial kits for applications in molecular biology:


HisExpress protein purification columns: http://www.claremontbio.com/HisExpress_Protein_Purification_Columns_s/72.htm (available at http://www.bioquote.com/product.php?id=291680)

PrepEase his-purification: http://www.affymetrix.com/estore/browse/brand/usb/product.jsp?navMode=34000&productId=131366&navAction=jump&aId=productsNav#1_1

Affinity chromatography products at Jena Bioscience: http://www.jenabioscience.com/cms/en/1/browse/1450_affinity_chromatography.html?gclid=CJCPmK6HvrUCFQtY3godFWEAtg


Kit Use Advantages Disadvantages Time Experience
Promega Wizard SV miniprep DNA isolation Quick Low yield 45 - 60 min
Promega PureYield™ Plasmid Miniprep SystemHigh-purity DNA isolation
Machery-Nagel NucleoBond Xtra Midi (midiprep) DNA isolation High yield (~20 ug), pure product. Slow 2 d Yields of 100-140 ng/uL in 200 uL, for a total of about 20 ug. Yield may depend on skill in handling washing and resuspension of DNA pellet. Might be more efficient to do several (~10) parallel minipreps instead?
Qiaquick gel extraction DNA extraction from gel Fast Impure product Impurities give absorbption peak at 230 nm in eluted product.
Qiaquick PCR purification Purification of PCR products Fast15 min
QIAEX II gel extraction DNA extraction from gel Higher purity than qiaquick Slower than Quiaquick1-5 h Yield of about 200 ng from ~1.5 ug starting amount of plasmid.
QIAGEN PCR Cloningplus KitSubcloning (TA cloning)?? ?
Dongsheng Bio Eco-friendly Plasmid miniprep kit DNA isolation  ? -
Dongsheng Bio PCR and DNA Fragment Purification KitPurification of PCR fragments  ?
QiaAmp


Notes:


Promega SV Wizard miniprep:

Observed plasmid yields:

Strain:Plasmid Date Culture volume incubated (mL) Culture volume centrifuged (mL) Medium /Antibiotic Incubation vessel Incubation time OD600 Elution volume (uL) Obtained DNA concentration (ng/uL) Total yield (ug) Yield (ug / mL * OD600) 260/280 260/230
Dh5a:rlig1 (pJP-1)21.5.1310 mL~4.5LB + Kan 50 ug/mL50 mL tube~ 8h~2.250 49 2,45 0,25
Dh5a:rlig1 (pJP-1)21.5.1310 mL~4.5LB + Kan 50 ug/mL50 mL tube~ 8h~2.250 35 1,75 0,18
Dh5a21.5.13 ~3.5LB 13 mL tube 2.7
Dh5a:pSB-M1g21.5.13 ~4LB + Kan 50 ug/mL 13 mL tube 2.6


DongSheng Bio Eco friendly plasmid extraction kit:


Observed plasmid yields:

Strain:Plasmid Date Culture volume incubated (mL) Culture volume centrifuged (mL) Medium /Antibiotic Incubation vessel Incubation time OD600 Elution volume (uL) Obtained DNA concentration (ng/uL) Total yield (ug) Yield (ug / mL * OD600) 260/280 260/230
Strain:Plasmid Date Culture vol incCulture vol centrmediumincub vesselincub timeOD600El. vol. conc tot rel 260/280 260/230


Qiaquick gel extraction kit:

Qiaex II: Use a thermomixer (high mixing, RPM ~ 500 ) to get constant mixing during incubation.


In vitro transcription kits:

https://www.neb.com/products/e2050-t7-quick-high-yield-rna-synthesis-kit

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