Where can we buy the enzymes needed for the LR recombination reaction?
You can order recombinase enzymes from Invitrogen.
Where can we get pDONR vectors?
You should order these pDONR vectors from Invitrogen. Some pDONR vectors are not in the Invitrogen catalog. However, Invitrogen is currently providing pDONR P3-P4 and pDONR P5-P6 as a custom service. The price is $500 USD for both vectors and they are only available as a pair. They provide 10 µg’s of DNA, maps and Vector NTI files for both vectors. To request these pDONR vectors, contact firstname.lastname@example.org.
It is also possible to order publicly-available pDONR vectors from GeneCorner
How can we design primers for N-terminal or C-terminal fusion to fluorescence tags?
What is the bacterial host strain of the plasmids?
The host strain of all plasmids is E. coli DB3.1 or ccdB survival cells from Invitrogen. Many plasmids contain the bacterial ccdB gene, which encodes an anti-DNA gyrase protein. The E. coli strain DB3.1 contains a mutation in the DNA gyrase gene so that the ccdB protein is unable to bind the mutant DNA Gyrase protein. In this strain the DNA is replicated normally and colonies grow.
DB3.1 can be acquired from GeneCorner.
Which antibiotic(s) can we use for growing the constructs?
The proper antibiotic is mentioned for all plasmids. The following concentrations are used:
Ampicillin (or Carbenicillin) 100 mg L-1
Kanamycin 50 mg L-1
Spectinomycin 100 mg L-1
Gentamicin 25 mg L-1
Chloramphenicol 30 mg L-1
Generally, binary vectors have spectinomycin resistance and high-copy plasmids are ampicillin resistant. Destination plasmids usually have a ccdB/CMR marker and so are also chloramphenicol resistant.
Why are my restriction digest/ sequencing results not compatible with the corresponding maps on this webpage?
We have sequenced most parts of the constructs, but unfortunately, there may still be some sequence errors in some vectors. Please do alert us if you find any discrepancies.
How are the vectors shipped?
Inside Europe, we send our parcels by first-class priority mail. Orders to destinations outside of Europe are sent by Federal Express.
How much does one vector cost?
The shipping and handling fee for each individual vector is 50 Euro. We recommend that you order at least 2 vectors for a minimum value of 100 Euro. You can order a single vector, but please note that this incurs a handling and shipping surcharge fee of €50. Starting from 2 vectors, this surcharge is waived. Belgian institutes and European Union institutes that have no VAT number must also pay 21% VAT.
How can I make sure my order will be allowed in by my country's customs?
For certain countries, we need additional data for customs clearance:
- Australia and New Zealand require an AQUIS import permit granted to your lab by the relevant government.
- Brazil requires a valid Brazillian company tax ID or CNPJ.
If your country requires this additional customs information, then please send it as soon as possible to the Gateway Vectors Shipping Staff.
I cannot see the VectorNTI files when I click on the VectorNTI link.
We cannot fully control how VNTI files are displayed by your web browser. To view the VNTI files you will need an installation of the Invitrogen VectorNTI software. Also, you or your system administrator need to configure your web browser properly for this to work. As a workaround, please download the VNTI files and import them into VNTI.