Smai cut site

Enzymes and Inhibitors.

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Smai cut site

Hi all, 1 Has anyone used smaI as a RE to cut a vector for blunt end cloning? So my situation now is I'm trying to cut a plasmid bp with smaI. I over digest overnight at room temp. Hope you guys can shed some light SmaI is not the best RE out there, but I understand that sometimes you'd have to use it since it's your only choice. I'd definately phosphatase my plasmid in order to prevent self ligation. I recently used Antarctic Phosphatase and it workes also very good. Good luck!!. Sma I has an isoschizomer which may be a better enzyme. If you still need the blunt end, you can just do a fill-in or chew back the overhang. Hi all, I juz started with cloning in pUC using SmaI to restrict digest for blunt ends and when i ran the gel got a linear plasmid but dint do dephosphorylation

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Listen to one of our scientific editorial team members read this article. Click here to access more audio articles or subscribe. Blunt ends can be easily created through PCR or enzymatic means. Blunt-end cloning is less efficient than traditional methods, and care should be given to avoid empty vector religation or insertion of fragments in both orientations. And do you know how to do both? Blunt and sticky might sound dull and dirty but knowing how these different cloning methods work is important when choosing which method to use. It is unlike sticky-end cloning, where both the insert and the vector contain single-stranded overhangs that complement each other. It would be challenging to perform partial digests that generate the sticky overhangs without also cutting up your PCR product Figure 1. Note that this is only true if you use a DNA polymerase with a proofreading function, such as Pfu.

Smai cut site

Enzymes and Inhibitors. Restriction Enzymes. Catalog number: ER Related applications: Restriction Enzyme Cloning. Technical Support Customer Service. Catalog Number. Save to list. Product Overview. See Reaction Conditions for Restriction Enzymes for a table of enzyme activity, conditions for double digestion, and heat inactivation for this and other restriction enzymes. Thermo Scientific conventional restriction endonucleases are a large collection of high quality restriction enzymes, optimized to work in one of the buffers of the Five Buffer System.

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Please try again or contact Customer Service. Search by lot number or partial lot number. Americas Brazil. There was an issue verifying your email address. We offer numerous convenient solutions to meet your lab's needs. Certificate of Analysis Search by lot number. For example, we may use these cookies to determine if you have interacted with a certain page. To prepare 10 mL of buffer, weigh 2. Preference Cookies We use these cookies to remember your settings and preferences. Hope you guys can shed some light To ensure consistent performance, Thermo Scientific restriction enzyme reaction buffers contain premixed BSA, which enhances the stability of many enzymes and binds contaminants that may be present in DNA preparations. Technical Support Customer Service. Log in with Your New Password. Contact Customer Service. Name This field is required.

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Technical Support Customer Service. This field is required. Failure to heat the water will result in PEG that will take approximately 24 hours to dissolve. If you still need the blunt end, you can just do a fill-in or chew back the overhang. For Research Use Only. Forgot Password? You can set your browser to block or alert you about these cookies, but some parts of our services will not work without them. This will prevent the Sma I ends of the vector from religating. For example, we may use these cookies to remember your language preferences. There was an issue creating your account. Your Account. Confirm Password Passwords don't match. A verified email address is required to access the full functionality of your Promega. Username already in use.