Transformation
To request a transformation you first need to fill in the form found under “request transformation” which you find here to the right. Please follow the guidelines stated there.
If you want to make a CRISPR CAS9 transformation we will treat that as 2 transformations which means we will have double amount of explants and you will also receive double amount of lines. Please try to put the name CRISPR in the name of the construct to avoid any misunderstanding. Regardless if you want a regular or CRISPR transformation. You need to provide the Agrobacterium containing the construct of interest as a pellet from a 50 ml liquid growing culture with an OD between 0.3-1.0 (max 1.5). The pellet must be provided in the morning of the given transformation day (preferably before 9.30 am).
The facility will then do:
- the transformation (regular 3 days), (CRISPR 3 days)
- selection on plates (Regular 2-3month),(CRISPR 2-6 month)
- growing the callus (Regular 1-1,5month), (CRISPR 1-2 month)
- growing the shoots (Regular 1.5-2month) (CRISPR 1,5-2month)
- selection of the shoots to get the individual lines (regular 1-2 month) (CRISPR 2-3 month)
If you forget your transformation appointment, we will charge for 1 hr work (400 SEK) (this is the time for us to prepare for the transformation in vain and putting everything back)
Selections available:
- Kanamycin
- Hygromycin
We can do transformations all year long but not during the summer (July-August) due to higher risk of infection.
Amplifications
To request an amplification, please fill in the form under the request tab to the right. Select “Amplification” and “New request”.
In the first table, fill in the database ID numbers of the transgenic constructs you wish to amplify. Add the genetic lines for each construct (one line per row) and the number of plants you want for each line. In the last column, select how many plants you want per jar when delivered. The default is three per jar unless stated otherwise.
In the next second table, add the wildtype you need for your experiment (T89 has database ID number 0 but it is also OK to write T89). Follow the same instructions as in the above section when adding details in the second table.).
Usually when starting an amplification, we only have 1-3 shoots to start with, so depending on how many replicates you want it might need several rounds of propagation. One propagation round normally takes between 4-5 weeks to grow. And most amplifications need several propagation rounds. So please plan. We will try to inform you when we start the amplification and when we expect it to be ready.
Taking samples for testing
We will inform you when your transgenic lines are ready to be tested. There are two possibilities:
- We can take the samples (stem or leave) for you as soon as the first individual lines are ready and freeze the samples in liquid nitrogen. The price for that is 35 SEK per sample
- The other possibility is to wait a few months more until the next routine cut and then you can get the old jars. The disadvantage of that is that we have less material to start with if you order an amplification of your selected lines.
For CRISPR transformation we take the samples by default in an earlier step so if you do not want samples you have to tell us in advance.