tol2kit.genetics.utah.edu
Basic Gateway principles - Tol2kit
http://tol2kit.genetics.utah.edu/index.php/Basic_Gateway_principles
NB: The Tol2kit uses three-insert multisite Gateway cloning (att4-att1-att2-att3) from Invitrogen. It is not. Easily compatible with the newer "Gateway Pro" system, for which the outermost sites are att1-att2. Principles of Gateway recombination. Here's a diagram of a Gateway BP reaction, showing an example of how a "middle" clone (pME) is constructed using PCR with gene-specific primers to add att. Sites, then recombination with a donor vector. How we use this in the Tol2kit. Given the different clones ...
tol2kit.genetics.utah.edu
List of entry and destination vectors - Tol2kit
http://tol2kit.genetics.utah.edu/index.php/List_of_entry_and_destination_vectors
List of entry and destination vectors. Here is a list of the constructs in the Tol2kit. Click on any construct name (second column) to see more information. 12/18/07: Clone pages now include annotated sequence in Genbank format. Are insert sequences for entry clones, and 5' and 3' ends for destination clones, along with a description of how to assemble predicted sequences for expression clones. Constructs in the Tol2kit v1.2 (Nov 2007). 5' entry clones, attL4-R1 (kan resistant). SV40 late polyA signal.
tol2kit.genetics.utah.edu
Tol2Kit:General disclaimer - Tol2kit
http://tol2kit.genetics.utah.edu/index.php/Tol2Kit:General_disclaimer
There is currently no text in this page, you can search for this page title. In other pages or edit this page. Retrieved from " http:/ tol2kit.genetics.utah.edu/index.php/Tol2Kit:General disclaimer. Log in / create account.
tol2kit.genetics.utah.edu
Protocols - Tol2kit
http://tol2kit.genetics.utah.edu/index.php/Protocols
Here are optimized methods and tips from the Chien lab, adopted after trial and error, which work consistently in our hands. See the Invitrogen multisite Gateway manual. For all of the basic information necessary to understand and perform Gateway recombination reactions. See also the Gateway tips. On the Lawson lab website. Please report problems or questions on the Tol2kit blog. What you will need. PCR Amplification of DNA. Purification of PCR products. Factors Affecting Reaction Efficiency. Provides ne...
tol2kit.genetics.utah.edu
Att site sequences - Tol2kit
http://tol2kit.genetics.utah.edu/index.php/Att_site_sequences
12-09-07 Minor edits to fix a couple of sequences and clarify names (especially the attBXX primer sequences). Att site shared sequences. Here is a set of "shared" sequences- the core stretch that is shared between attB/P/L/R for att1-4. These are useful for stitching together entry and destination sequences to calculate the sequence of a final expression clone. Att1 shared TTTGTACAAAAAAG att2 shared CTTTCTTGTACAAAGT att3 shared CAACTTTATTATACA att4 shared CAACTTTGTATAGAAAAGTTG. List of att sites.
tol2kit.genetics.utah.edu
P5E-UAS - Tol2kit
http://tol2kit.genetics.utah.edu/index.php/Special:Random
P5E-UAS was made by conventionally subcloning into p5E-MCS a HindIII-EcoRI fragment from pBUAS-E1b-RFP from Reinhard Köster, containing multimerized Gal4 upstream activating sequence (UAS) elements, followed by an adenovirus E1b TATA box and then by a piece of the carp beta-actin 5' UTR. Functional tests of constructs using this element show strong activation by Gal4. Annotated sequence, Genbank format:. FASTA file with the full-length sequence as well as sequences of individual components:.
tol2kit.genetics.utah.edu
Sample results with the Tol2kit - Tol2kit
http://tol2kit.genetics.utah.edu/index.php/Sample_results_with_the_Tol2kit
Sample results with the Tol2kit. Here are some examples of results obtained with the Tol2kit. They are included in a methods paper that has recently (September 2007) been accepted at Developmental Dynamics. Validation of IRES clones. If you are interested in testing these constructs, please contact Chi-Bin Chien. We also encourage others to generate other IRES constructs, for instance using CFP or red fluorescent proteins such as DsRedExpress or tdTomato. Log in / create account.
tol2kit.genetics.utah.edu
Tol2Kit:Community Portal - Tol2kit
http://tol2kit.genetics.utah.edu/index.php/Tol2Kit:Community_Portal
There is currently no text in this page, you can search for this page title. In other pages or edit this page. Retrieved from " http:/ tol2kit.genetics.utah.edu/index.php/Tol2Kit:Community Portal. Log in / create account.
tol2kit.genetics.utah.edu
Main Page - Tol2kit
http://tol2kit.genetics.utah.edu/index.php/Special:Whatlinkshere/Main_Page
No pages link to Main Page. Retrieved from " http:/ tol2kit.genetics.utah.edu/index.php/Special:Whatlinkshere. Log in / create account.
