CLONES & CLONING SERVICE
What is the difference between MGC and ORFeome libraries?
- In ORFeome library all clones are ORFs in gateway compatible entry-vectors (pENTR221, pENTR201, pENTR223)
- In MGC library the clones are cDNAs in different vector backbones. Check the vectors from the library excel.
For more information please see:
How do I know whether you have the clone I'm interested in?
- Check the excels from the Cloning page.
What if I can't find my clone of interest?
- You can check the websites of commercial providers e.g. invitrogen, addgene, genecopoeia (check their validation status for the clones).
- If the clone you are looking for cannot be found in the excel sheets, it is unfortunately not included in our collections. However, make sure you search with all possible gene names and acronyms and please note that there are separate sheets for human and mouse constructs.
- If your clone of interest can only be found in the MGC collection but the vector is not suitable for your experiment, it is possible to produce a Gateway entry clone by PCR (this can also be applied if you have the cDNA yourself in any vector). Please ask for more details regarding this customized service!
How can I make a clone request?
- Please fill in the clone request form and send it to us as an email attachment. Remember to add your research group, clone name and position and possible cloning service needed.
What information is available for my clone?
- The ORFeome library is validated by the vendor and we have performed random validations by sequencing. MGC library is validated by the vendor by sequencing the N-terminus of the clones.
- For checking clone sequences, search database with the accession number stated for each clone in the excel (in most cases the accession number has prefix "BC").
Should I pay attention to the stop codon status of ORFeome entry clones?
- The ORFeome collection contains both entry clones with stop codon and entry clones without stop codon. The stop codon status is stated for each clone in the excel sheet.
- If you'd like to request a cloning into an expression vector where your protein would be expressed with a C-terminal tag/epitope fusion, you need to choose an entry construct without stop codon for the cloning.
Do I need stable E. Coli cells for growing lenti constructs?
- It is recommended to grow lenti constructs in E. Coli (Stbl3) as the lentiviral DNA contains direct repeats and is less stable.
- If you need to inoculate new cultures for preparing more lentiviral DNA, transform the constructs to Stbl3 cells or use the glycerol stock obtained from GBU. Please ask for the glycerol stock at the same time you receive the constructs.
How do I get my clones?
- We provide the clones as a streak on a bacterial plate and cloned ORFs as a miniprep. We send plates/tubes by mail or you can pick them up. If requested, you can also obtain the glycerol stock when picking up the plate/miniprep, but we don't send glycerol stocks by mail.
Should I sequence my clones?
- We highly recommend sequencing the clones before using them in experiments.
- Suitable sequencing primers are stated in an info sheet you receive as email attachment when your order is ready.
- Viikki sequencing service and Biomedicum sequencing unit have primers for sequencing constructs from GBU.
What if there is an error with a clone I receive?
- The clone collections are sequence verified by the vendor. GBU has validated the libraries by random sequencing and observed some errors on the plates. On this account we highly recommend sequencing the clones before using them in experiments.
- If sequencing shows the clone you have received is not the one you requested, please contact GBU immediately. Please make sure you compare the sequencing result to the exact same sequence that is indicated in the excel sheet for your clone.
- Upon your notification on the wrong clone we will check the plate it originated from to rule out errors with picking the clone.
- If possible, incorrect clones are reimbursed by picking the same or equivalent clone from another location in the library.
- If the same or equivalent clone cannot be found in other locations in the collection, the incorrect clone will not be charged but cannot be compensated otherwise.
Can I request empty vectors from GBU e.g. for transfection controls?
- GBU does not provide empty Gateway destination vectors.
- If you need a control construct we have enhanced GFP and mCherry sequences cloned into destination vectors. Please indicate the need for a control construct in the clone request form.
Does the cloning service include the cloning of customers cDNAs and/or to customers vectors?
- Please send a request via email if you are interested in using your own constructs in cloning.
Please note: GBU clones are under MTA terms from the suppliers. See MTA terms provided with a clone. The fee is solely from transportation and packaging.
DIGITAL SLIDE SCANNING SERVICE
How do I know if my slides are suitable for scanning?
- The scanner takes standard 25x75mm microscope slides and allows variation in slide thickness between 0.9-1.2mm. If you are not sure whether your slides meet these requirements GBU has a tool for measuring the slides.
- All slides to be scanned must be mounted with a solidifying mounting media and the coverslips must not protrude over the edges of the microscope slide (see more instructions for prepping the slides below).
- For each sample type we can perform a test scanning free of charge.
Do I need to know how to operate the equipment and how can I make a reservation?
- Slide scanning is operated as a service, so GBU will take care of the scanner runs and provide you with the images. However, we would be happy to do test scannings with you as well as demonstrate how the equipment works if you are not familiar with this technique.
- For reservations: please send email to firstname.lastname@example.org describing the type of samples you have and how many slides you'd like to have scanned. GBU will suggest a schedule for your scanning project depending on the current situation.
What kind of image files the scanner produces and how can I view them?
- The image data for each slide consists of a folder and a .mrxs-file with the same name. When copying the images, please make sure you copy both the folder and the .mrxs-file in order to open the image in the viewing software.
- You can view the slides using a 3DHISTECH software (Pannoramic Viewer or CaseViewer, both can be freely downloaded from their website). The softwares are available for Windows only. You can also view your images using Biomedicum Imaging Unit imaging workstations.
- Using the viewing softwares it is possible to export selected areas in tiff or jpeg format.
How do I receive the image files?
- Please note that the image files can be large (up to several GBs / slide with bright field). Images are saved on the scanner computer R: drive immediately following scanning. The network connections of the scanner computer are restricted and for the brightfield images it is recommended you copy the files straight from the computer to your own portable drive. As the fluorescent images are usually smaller, it is possible also to distribute them using Funet FileSender server.
- You can copy the images while the scanner is running. Please do not remove the files from R: drive, just copy them to your own disk.
Can I have the image files named according to my samples?
- If the scanner has generated names to your image files they are named with a running number from 1M01 to 1M25 for the first magazine capable of taking 25 slides, then 2M01-2M25 and so forth.
- For the automated bright field runs it is possible to import sample names to the scanner and the image files will be named accordingly. Please ask for more details on how to provide GBU with the name list.
- It is also possible to change the image file names later on utilizing standard file management. However, please make sure you rename both the folder and the .mrxs-file.
- The scanner is equipped with a bar code reader if your slides are labeled with 1D or 2D barcodes.
How can I see the frosted end of the scanned slide?
- It is always possible to see the frosted end of the slide with the sample details. In Pannoramic Viewer when viewing one slide, click the button with "i" icon on top ot the window. Choosing "slide information" will open a window where with an image of the frosted end and details of the scanning (objective, scanning time, image file size, layers, for fluorescence exposure times).
What if I receive an image that is not focused well, parts of the sample is missing or is otherwise of poor quality?
- Please contact GBU for rescanning the slide. For difficult samples it is possible to use more focusing points or apply extended focus option.