Chamberlin Lab Protocol Guide (archive)
A collection of lab protocols and general lab practice guide on molecular biology from Chamberlin lab at Ohio state University.
Content:
General Lab Procedures
Autoclaving .. 4
Disposal
Disposal of Burn Boxes .. 4
DNA Strider 4
Getting Reagents on Site
Dean Lab 5
Fermentas Freezer .. 5
Ostrowski Lab (Qiagen Products) .. 5
Mailing Parcels 5
Making a Worm Pick . 5
Ordering
Ordering Primers 6
Sequencing (at Neurbiotechnology Center) . 6
Sterilizing
Washing Glassware/The Glassware Facility... 7
Centrifuge Tubes 7
Double Distilled Water ... 7
Pipets (Glass) .. 7
Plastic Tubes ... 8
Test Tubes (Glass) .. 8
Tips . 8
Toothpicks ...... 8
Streaking Plates .. 9
Worm Plates 9
Removing Contaminants from a Plate 9
General Lab Tips
Emergencies 10
Sterile (Aseptic) Technique ... 10
Pipetting .. 11
Using Equipment 11
Lab Protocols
Competent Cells
Making Competent Cells ... 11
Transforming Competent Cells (to check them).. 12
Checking Cell Competency ... 12
Cosmids, Receiving 13
DIG (Non-Radioactive Labeling) ..14
Semi-quantitative determination of labeling
Efficiency. 14
Hybridization 17
Immunological Detection . 20
Striping and Reprobing DNA Blots . 21
DNA Quantification
Spectrophotometry ... 22
Ethidium Gel Plates .. 22
Enzyme Reactions 23
Freezing Stocks
Making Frozen Stocks . 24
Thawing Frozen Stocks 28
Gels
DNA Gels . 30
Gel Band Cutting . 34
Gel Band Purification .. 35
Images
Developing X-ray Film 37
Exposing X-ray Film 37
Inoculating LB (LB + Op) ... 38
Ligation Reactions 39
Liquid Cultures . 40
PCR
PCR . 41
PCR Purification . 46
Worm Lysis/PCR 70
Plates
LB/LB antibiotic plates ... 47
Pouring Nematode Growth Media Plates ... 48
Spreading NGM Plates ... 49
Preps
Alkaline Lysis (Mini Prep) .. 50
Qiagen Plasmid Midi Prep .. 52
Qiagen Spin Column Mini Prep .. 55
Solutions
CaCl2 (1.0M, 1.0 L) ..57
EDTA (0.5M, pH 8.0, 1.0 L) ... 57
Freeze Solution with agar (500 ml) . 58
Glycerol Solutions ... 59
LB Media (2.0 L) . 59
LB Media with MgSO4 and Maltose (500 ml) 60
MgCl2 . 61
M9 Buffer (1.0 L) 61
NGM (Nematode Growth Medium) (2.5 L) 62
P1 Buffer (Resuspension Buffer) (500 ml) . 62
P2 Buffer (Lysis Buffer) (500 ml) .. 63
PBS (Phosphate-Buffered Saline) pH 7.4, 1.0 L . 63
Potassium Ferricyanide (100mM, 10 ml) ... 64
Potassium Ferrocyanide (100mM, 10ml) 64
PPB (KPO4 Buffer or Phosphate Buffer)
(1.0M, pH 6.0, 1.0 L). 65
Proteinase K (10 mg/ml from 50 mg/ml stock) ... 65
SM Buffer (1.0 L) 66
Sodium Acetate (3.0M, pH 5.2, 500 ml) . 66
Tris Cl Buffer (1.0M, pH 8.0 or other) . 67
Worm Lysis Buffer .. 68
Transformation 69
Worm Lysis .. 70
REFERENCES 71
Last update 03-May-2005, Rating n/a of 0 votes.
