Transformation: 4500 SEK
Includes transformation and propagation of about 10-14 individual lines (“1st mother Jar”)

Routine cut:
Internal: 60 SEK / Jar
External: 100 SEK / Jar

Amplification:
Internal: 60 SEK / Jar
External: 100 SEK / Jar

Propagation of T89, Elite 864012 or Tobacco:
Internal: 75 SEK / plant
External: 100 SEK / plant

Routine cut after 3 years:
150 SEK / Jar

If the transformation fails:
One retransformation is offered for free.

Forgotten transformation: 500 SEK
(We still have to prepare and we lose at least one hour work)

Cut in 50 ml falcon tube for transport:
Internal: 60 SEK
External: 100 SEK

Samples:
35 SEK / Sample (tube)

Jars:

Empty MS2 jar 10 SEK, Empty MS6 jar 10 SEK
Filled MS2 jar 60SEK, Filled MS6 jar 35 SEK

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 can give us the plasmid, and we will make the agrobacterium transformation, or you can provide us with 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 ( before 9.30 am).

The facility will then do:

• the transformation in agrobacterium (1 week)
• the transformationin poplar (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 (500 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:

1. 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
2. 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.

Use your normal UMU-ID or SLU-ID login to access the request form

Procedure for transformation request:

• Fill in the request form and submit new request through the link requests.
• Choose the construct name carefully, so that it is possible to identify the construct in the database in the future, both for yourself and for others. It is not possible to change the name later when we have started the transformation.
• Your transformation will be assigned a database ID number by lab manager Veronica Bourquin.
• The platform will notify you when your request is scheduled for transformation minimum 1 week in advance, so that you can prepare the bacteria culture.
• One member of our team will be assigned to perform the transformation and the following cultivation of your construct.

Note! The requests will be handled in the order they are submitted to the platform.

Good to know prior to the day of transformation:

• Bring your Agro pellet at 09:00-09:30 on the day of transformation (we do not accept arrivals later than 10:00!)
• Prepare the pellet from a 50 ml growing culture with an OD₆₀₀ of 0.5-0.8 (no lower than 0,3 and no higher than 1.5!)

After the transformation event, we will update you when it is time to sample and genotype the lines.The poplar transformation procedure: 2-6 months - Infection with Agrobacterium and selection of transgenes on plates; 1.5-2 months - Induction of shoot elongation in transgenic calli; 2-3 months - Transfer of shoots to new medium for root formation; 1-1.5 months: Transformation completed - the "1st Mother jar" is ready

Procedure for requesting poplar transformation:

Access the request form through the link above Request Amplification - New Request, Good to know prior to fill in the amplification request.

• Use the construct ID number generated before the transformation and state which lines you want (one line per row)
  If you do not have the construct ID number, use the construct name originally submitted for the transformation and write that in the comments and we can help you find out the correct ID number!
• Amount of plants
  We always propagate extra (approx. 10%) to increase the chance to meet the amount requested. (Write in the comments if you do not want extra plants).
• Plants/jar: there will be three plants in each jar by default.
• Wallenberg Greenhouse experiment: Announce whether the amplified lines are planned for potting in the Wallenberg Greenhouse (YES/NO). Note that this is not a booking! Please contact This email address is being protected from spambots. You need JavaScript enabled to view it. for booking space at Wallenberg, in climate chambers at UPSC floor 5 or at Skogis.
• Homogenous (Yes/No), is homogeneity important for your experiment?

Note! Amplifications take time. For example, an amplification of 15 replicates generally needs 2-3 propagation rounds. First propagation takes at least 5 weeks to grow, the following takes about 4 weeks each. If you want them more homogeneous, 1 or 2 extra propagations may be needed. Also, if you want less than 3 plants in each jar by delivery it will need an extra propagation round.

First cut is a kick start and takes about 4-6 weeks, then depending of how many replicates it need 1-3 more amplifications rounds. When we have the final number of plants we can make the fianl cut and then the amplification is ready usually between 8-27 weeks

Reverse genetic approaches are the only realistic alternative for the analysis of gene functions in trees because of their long generation time and obligatory out-crossing of most trees. UPSC scientists were among the first in the world to be able to create transgenic trees.

Poplar in vitro room 

All initial work was done with a fast-growing and relatively easily transformable hybrid aspen (Populus tremula x tremuloides) clone originating from the Czech Republic called “T89”. This is still the main tree species used in the transformation facility, but the numbers of different tree species that are routinely transformed have now grown to include elite varieties of hybrid aspen, Populus trichocarpa, Populus alba, Populus tremula and also Betula pendula (silver birch).

The method used is Agrobacterium-mediated plant transformation of stem segments, petioles or leaf explants and the facility is today growing more than 15 000 transgenic trees, representing more than 5000 different lines, probably making this the largest tree transformation facility in the world. The transformation facility currently employs five fulltime persons and is servicing all the research groups at UPSC and collaborators at both Umeå University and the Swedish University of Agricultural Sciences. Any group requiring transformed trees simply needs to provide the Agrobacterium strain carrying the genetic construct they want to analyze. After 8-10 months the trees are ready to be transferred to the greenhouse for analysis.

Different stages of poplar transformation: from explants to small trees on soil.

The persons working at the facility:

Contact:

For practical questions e.g. regarding the transformation procedure, please contact:
Veronica Bourquin at This email address is being protected from spambots. You need JavaScript enabled to view it.

For other questions, please contact:
Ove Nilsson at This email address is being protected from spambots. You need JavaScript enabled to view it.