DNA实验技术

Procedure:1) Need to cut 10 µg of plasmid in two spots (‘A’ and ‘B’) in polylinker. ‘A’ cut is with a restriction enzyme which gives a 3’ recessed end (exonuclease sensitive) next to insert. Since ...

0.25M HCl (1L)20.66mL HCl 979.34mL dH2O *add acid to wateundefined 1M ammonium acetate 0.02M NaOH (1L)77.08g ammonium acetate 0.8g NaOH Fill with dH2O 0.5X SSC 0.1% SDS (1L)25mL 20X SSC 10mL 10% SDS 965mL dH ...

Smith and Summers 1980. Anal. Biochem. 109:123-129.Digest 10-15 µg genomic DNA with desired restriction enzyme overnight at 37oC. Run digest on a 1% TBE agarose gel at 100V until 1st blue dye reaches ...

Genomic DNA Digest10 µl DNA (10 µg) 3 µl 10X Buffer 1-2 µl Restriction Enzyme (40 units) 3 µl 10mM Spermidine-HCl 3 µl 1 mg/ml BSA 10 µl H2O Total volume: 30 µlNote: Use DNA from a standard tail prep ...

SOLUTIONSLysis Buffer 0.1 M Tris pH 8.0 0.2 M NaCl 5 mM EDTA

Principle: Refer to Southern Transfer with Zetabind membrane. The NaOH transfer may be a more efficient transfer method for larger sized fragments. It is the method of choice for pulsed-field gels con ...

I. Preparing worm genomic DNA: requires 1-2 days to seed agarose plates a few days for the worms to grow 1-2 days to prep the DNA1. Seed large agarose plates with HB101. Agarose is preferred over agar ...

MaterialsHot Probe dNTP Mix:25 mM dATP25 mM dTTP25 mM dGTP2.5 mM dCTPHybridization Solution:5X SSC0.5% (w/v) Blocking Reagent0.1% (w/v) N-lauroylsarcosine Na-salt0.02% (w/v) SDS50% FormamideBlot Wash ...

IRS-1 Probe DNA Prep 1. From glycerol stocks (in �80ºC freezer) grow up bacteria. Use amp LB. 2. Use a Qiagen prep to purify the DNA. 3. Perform the following digest: ...

Preface: Digesting lambda DNA (BRL) with a specific restriction enzyme will result in lambda DNA fragments of known size. Three separate lambda digests (BglII BstEII and XhoI) will give 23 fragments w ...

You will require the following: -Tail buffer 50ml 10% SDS 5ml 1M Tris pH 7.50.5ml 0.5M EDTA5ml 5M NaCl 1.5ml DDW38ml -Phenol/chloroform (1:1 mixture) -0.5M EDTA -4M NH4Ac -Absolute EtOH -70% EtOH -T ...

About 1/2 to 1 cm of tail should be cut from properly marked mice (using toe or ear clipping etc) about the age of 2-3 weeks. Place these tails into marked 1.5 ml Eppindorf tubes for processing; they ...

About 1/2 to 1 cm of tail should be cut from properly marked mice (using toe or ear clipping etc) about the age of 2-3 weeks. Place these tails into marked 1.5 ml Eppindorf tubes for processing; they ...

A. Large scale double-stranded DNA isolationThe method used for the isolation of large scale cosmid and plasmid DNA is an unpublished modification (16) of an alkaline lysis procedure (1718) followed b ...

We store our all of our plasmids as bacterial host stocks at -80 stored in 7% DMSO. Such stocks are very long-lived giving robust bacterial growth even after 10-15 years. However occasionally we hav ...

1) Grow 3-5 ml over-night E. coli cultures containing your plasmid2) Spin down 1.5 ml cells3) Resuspend in 300 µl Buffer P1 w. RNase A4) Add 300 µl Buffer P2.Mix by invertin ...

Stuff you need:TENS Buffer10 mM Tris-HCl pH 8.01 mM EDTA pH 8.00.1 N NaOH0.5% SDSTE pH 8.010 mM Tris-HCl pH 8.01 mM EDTA pH 8.095% EtOH precooled to -20 deg C70% EtOH at room temperature3 M Na Acetate ...

Objective:Mini-Prep procedure is used to isolate small plasmid DNA from bacteria while limiting contaminating proteins and genomic DNA. The plasmid quality is acceptable for restriction analysis seque ...

PREPARE SOLUTIONS 1. Glycerol mix (1 L):Weigh 25 grams (liquid weight) of glycerol and add dH2O to 1 Liter (Autoclave)2. Potassium mix (1 L):Mix 170 mL of 1M KH2PO4 (monobasic) 720 mL of 1M K2HPO4 (di ...

1. Shake E. coli harboring plasmid at 37 C overnight in 50 ml of TB containing appropriate antibiotics. (when using ampicillin addition of the antibiotics to 100-200 ug/ml rather than usual 50 ug/ml m ...