Frequently asked questions from our customers
Questions about Target Genotyping
For the establishment and validation of each target we need information about the primer sequences used, the size of the PCR products, and - if available - JAX number and the DNA sequence of the construct. The latter is only required in very rare, complicated cases. If a 3-primer system is used, the information about which of them is the universal primer that is always involved in the PCR with the other two is very helpful. In the appendix you will find our Target sample sheet, in which all required information can be entered.
We need reference material for the validation to test the primers. Ideally, this would be a heterozygous sample (wt/mut) and a homozygous one (mut/mut). Of course, the latter is only possible for transgenes whose integration site into the genome is known. Reference samples should always be shipped in well-labeled, individual 1.5 ml reaction tubes. A wt/wt reference is generally not required, as any sample can be used for this purpose.
Since we use a DNA sequencer for genotyping, which works with fluorescently labeled primers, we always perform our own primer design. We are limited to a length of the PCR products in the range of 100 bp to 500 bp. In most cases, the customer's original primer sequences can be taken, as long as the specific PCR products are not larger than 500 bp. Otherwise, we have to try out different primer variants on suitable reference material.
You can either send biopsy material, such as tails, ear punches (one ear punch is sufficient) or sample material on swabs.
Ready extracted DNA can also be sent. If possible, do not use "quick-and-dirty" extractions, as they do not necessarily harmonize with the PCR chemistry we use. If you are not sure, contact GVG Genetic Monitoring and have your DNA extraction protocol checked. We recommend to send about half of the DNA extract to us, the other half remains with you as a reserve sample.
Biological material is perishable. Such material is known to degrade within a few hours at room temperature, whereas it can be stored refrigerated for a longer period, ideally in a frozen state. Therefore, it is important that the cold chain is not interrupted after sample collection and for shipment. Ideally, sample material should be shipped at the beginning of a week. This ensures that the material is not stored unrefrigerated over the weekend.
We guarantee the transmission of the test results within 10 working days after receipt of the sample. However, experience shows that the period is considerably shorter.
Unlike many competitors who only submit the PCR results, we perform the full genotyping and assign the exact genotype to the sample based on this. This allows the customer an automated upload of the results directly into the management software for breeding established in the research facility. This clearly sets our service apart from the competition.
The genotyping results are transmitted as a csv file. You can import our analysis data directly into your management software without own writing effort, e.g. into PyRAT. The company Scionics from Dresden, which provides the software PyRAT, has created an extra interface for the GVG, with the help of which the analysis data delivered by us as csv file are directly uploaded into PyRAT and assigned to the individual mice. This saves a lot of manual work on your part and avoids transmission errors! If you use another software, please contact GVG in time, so that a solution can be created together with the provider of the software.
However, the automated upload only works if the nomenclature of the genotypes in the csv file is 100% identical to the information in the database. We take all this information from the target sample sheet.
- the exact spelling of the mutation name (including upper and lower case)
- the exact designation of the different alleles (wild type: wt or +; mutant: fl, mut, tg, etc.)
- the order of entry of the alleles: wild type first or the mutant. This order must be consistent for all targets
We calculate the costs for the genotyping of a sample based on the number of targets to be analyzed. It does not matter whether only one primer pair is needed for the detection, or 3 or 4 primers (e.g. 2 primers for the wild type and 2 others for the mutant). If two different, three or more targets are genotyped in parallel for one sample, the price for the second target and the subsequent targets is reduced.