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... of cold 4% paraformaldehyde in PBS on ice for 15 min . Pro-Tip While 4% Paraformaldehyde fixation works...rocking platform. Permeabilize cells for 10 min at room temperature ( RT ) on a rocking platform in 500...rocking platform. Section 3: Labeling with antibody Block for 20 min at RT on a rocking platform in 500...labeling HeLa cells for a target protein using the formaldehyde fixation method. The protocol may need to be...specific products or equipment. Inclusion of this information is solely for transparency intended to support...Pipette controller Pipette tips and pipettes Rocking platform Tweezers Fluorescent microscope 0.5–10 µL single...coated coverslips HeLa cells 24-well plate 4% Paraformaldehyde 5 mg/mL 4′,6-diamidino-2-phenylindole (DAPI...

...incubation, transform DH5α E. coli cells by heatshock with the plasmid of interest. See our transformation page...for a detailed heatshock transformation protocol. Plate 50 µL of transformed E. coli /rescue media suspension...also like... Making LB Agar Plates Bacterial Transformation Recovering Plasmid DNA from Bacterial Culture...plating antibiotic over-agar for the selection of transformed E. coli . Over-agar spreading of antibiotic makes...investigator to conveniently plate and select transformed cells containing plasmids differing in their ... sterile water (or other antibiotic) E. coli transformed with a plasmid containing the carbenicillin (...for 18 hours. Day 2 Observe plates for colony formation. Shown below are the results from an experiment...

... a shaking platform. Wash the membrane 3x for 5 min in 1X TBST at RT on a shaking platform. Section 6:... a shaking platform. Wash the membrane 3x for 5 min in 1X TBST at RT on a shaking platform. Prepare the...specific products or equipment. Inclusion of this information is solely for transparency intended to support...supply iBlot 2 Gel Transfer Device Roller Spatula Platform shaker Cold room Gel imager -80 °C freezer Reagents...the manufacturer's instructions for additional information specific to your antibody, such as ideal blocking...between antibodies. Refer to the manufacturer's information before running your experiment. Remove the membrane...overnight in primary antibody at 4 °C on a rocking platform. Pro-Tip Primary antibody incubation can be reduced...

...self-ligating recipient plasmid backbone. Transformation Transform your ligation reaction into your bacterial...from your transformation will give you the first indication as to whether your transformation worked. our...Information about plasmid cloning by restriction enzyme digest (subcloning), including design and experimental...Digest of Plasmid DNA DNA Ligation Bacterial Transformation Summary The following technique can be used...in a new window) New England Biolabs for more information about restriction enzyme buffers). If you select...competent cells. For most standard cloning, you can transform 1-2μl of your ligation reaction into competent...conduct a positive control to ensure that your transformation worked. You should also verify that you are...

...Digest of Plasmid DNA Bacterial Transformation Background Information The final step in the construction...ligation, is performed by the T4 DNA ligase enzyme. The DNA ligase catalyzes the formation of covalent ...the backbone and the complete plasmid can be transformed into bacterial cells for propagation. The majority...digested vector backbone with a phosphatase before performing the ligation reaction (phosphatase removes the... incubation at 37°C. Proceed with bacterial transformation . Tips and FAQ Do controls When doing ligations...contamination of intact plasmid in ligation or transformation reagents Optimizing the Vector:Insert Ratio...

...addgene.org to get started. Genes KLF gene pages with information on available plasmids, references, and links....

...academic and industrial organizations. For more information on ordering from a for-profit entity, please ...

...standards to study protein-protein interactions or conformational changes within a protein. Biosensors Monitor...

...Esposito Lab for use with the Gateway® Cloning Platform (Life Technologies™) to permit construction of...

...process online. Click here for additional ordering information....

...all pX458 family plasmids. Additional Cloning Information Cloning Protocols Single guide cloning: Zhang...

...Lab plasmids Scot Wolfe Lab plasmids Detailed Information Protocols & Resources Contents of Kit Original...

...specific products or equipment. Inclusion of this information is solely for transparency intended to support...Microcentrifuge Electrophoresis chamber Power supply Rocking platform Fume hood Metal spatula Razor blade Plastic tray...water for 5 min with gentle agitation on a rocking platform. Pour off the water in the sink. Add 20 mL of ...incubate for 1 h with gentle agitation on a rocking platform. Pour off the SimplyBlue SafeStain in the sink...incubate for 1 h with gentle agitation on a rocking platform. Pour off the water in the sink. Add 100 mL of...incubate for 1 h with gentle agitation on a rocking platform. Pour off the water in the sink. Take a brightfield... exported CSV file. Select import. Choose file. Format as needed. Select the Insert tab, then Chart . ...

...step and proceed to binding of RISC. For more information on shRNA design and delivery, see resources below...

... Bio (Link opens in a new window) . For more information about PRISM, see the preprint: Park, S.Y., Sheridan...

...promoter, and so on. The following table provides information for the assembly standards most commonly used...

...Bacterial Transformation Recovering Plasmid DNA from Bacterial Culture Background Information Bacterial... not need to obtain more competent cells and retransform. Bacteria on an LB agar plate can be stored at... temperature, can be found on your plasmid's information page. The next day you will be able to start ...freezing (5-6 times). Make sure that you see one uniform solution, and there are no layers present. Be sure...

...Follow this protocol to perform RNA extraction without a kit....Water-saturated Phenol 2 M Sodium Acetate pH 4 Chloroform/Isoamyl alcohol (49:1) 75% Ethanol RNase-free...phenol, mix thoroughly by inversion. Add 0.2 mL of Chloroform/Isoamyl alcohol (49:1) and then shake vigorously...Nanodrop), agarose gel, or bioanalyzer. For more information on nucleic acid quantification, see our protocol...from the homogenized sample(s). Add 0.2 mL of Chloroform/Isoamyl alcohol (49:1) per 1 mL of TRIzol® used...

...Tagging in Human Cells Find additional cell line information from the Allen Institute for Cell Science data...

...arrays in a 2012 issue of Nature Protocols . More information and help can be found at TALeffector Resources...