Server at Davidson College
Instructions:
Plasmid/Gene sequences:
Paste or enter the single strand sequences of the plasmid and other components you are working with in FASTA format.
Enter the sequences in the 5' to 3' direction, with new sequences on separate lines.
The sequences will be screened for restriction enzyme sites.
Restriction enzyme:
Select from menu the restriction enzyme you will be using.
This will change the sticky ends that the generator uses as well as the cutting behavior of the enzyme.
Melting temperature:
Enter the desired melting temperature of the junction. Note that there is a minimum melting temperature, which will depend on junction sequence length. For junctions 18 base-pairs long, the minimum melting temperature is 44 °C. If you are having difficulty finding junctions, it is possible that your melting temperature is too low, which results in a large proportion of A and T in your sequence. Try raising melting temperature and re-submit. GGAJET uses the following equation to determine melting temperature: 
Junction length:
Enter the desired length of the junctions in base pairs.
Number of desired junctions:
Enter the number of junctions you will need. Note that more junctions will require more run-time.
PCR Annealing temperature:
Enter the annealing temperature you will use, in degrees Celcius. Note that this field is optional.
If this field is left blank, GGAJET will assume an annealing temperature of 5 °C below the melting temperature.
Junction Testing:
Add one junction at a time and test against the plasmid. You can add a maximum of 10 junctions. Do not enter the reverse complement; it will be added automatically.
"EMBOSS: The European Molecular Biology Open Software Suite" (2000)
Rice,P. Longden,I. and Bleasby,A.
Trends in Genetics 16, (6) pp276--27
PubMed: 10827456
DOI: 10.1016/S0168-9525(00)02024-27
"RNAstructure: software for RNA secondary structure prediction and analysis."
J.S. Reuter and D.H. Mathews.
BMC Bioinformatics, 11:129. (2010).
PMCID: PMC2984261
"IDT SciTools: a suite for analysis and design of nucleic acid oligomers"
Richard Owczarzy, Andrey V. Tataurov, Yihe Wu, Jeffrey A. Manthey, Kyle A. McQuisten, Hakeem G. Almabrazi, Kent F. Pedersen, Yuan Lin, Justin Garretson, Neil O. McEntaggart, Chris A. Sailor, Robert B. Dawson, Andrew S. Peek
Nucleic Acids Res. 2008 July 1; 36(Web Server issue): W163–W169. Published online 2008 April 25.
doi: 10.1093/nar/gkn198
PMCID: PMC2447751
The GGAJET (pronounced "gadget") was designed by Jackson Spell and Micah Brown as a part of the 2013 Davidson College/Missouri Western Synthetic Biology Summer Research Project, under Dr. Laurie Heyer and Dr. A. Malcolm Campbell. This was made possible through the NSF MCB 1120558 and NSF MCB 1120578 grants.
Please send comments, questions, and suggestions to Laurie Heyer at laheyer@davidson.edu
GGAJET was produced as a research tool and is not intended for commercial purposes. Davidson College assumes no responsibility in relation to the use of GGAJET or any information produced by it.
