Molecular genetics: mouse genotyping

Delivering excellence in rodent genotyping to researchers nationally

About
Our services
Accessing our services

Garvan Molecular Genetics (GMG) offers a rodent genotyping service for mice and rats. DNA is extracted from submitted tissue samples and extracted via our high-throughput robotic platforms. PCR is set up with fully automated liquid handling systems that are controlled by our in-house developed programming software.

We use a combination of realtime PCR and high-resolution melt curve analysis to deduct the genotypes, similar to Jackson Laboratories in the US. Results are sent to clients via email. DNA samples are stored for six months and can be sent to clients for further experiments upon request.

We are committed to providing an excellent service quality. Therefore, we continuously improve our services to achieve highly reliable results, short turnaround time and minimal costs. Client satisfaction is monitored by annual surveys and constant feedback.

Sample DNA quality

The analysis method of our Mouse Genotyping protocol is sensitive to the quality of the DNA that we are given. This is due to the fact that we run a realtime PCR with a program suitable for most primer annealing temperatures. We have observed that purified DNA gives the best results.

If you send us DNA samples rather than tissue, we recommend cleaning the proteinase K digestion via columns or magnetic beads. Any Qiagen or similar clean-up kit will work effectively. We will accept DNA that has not been treated in the recommended way but we must make you aware that we observe a dropout rate of ~20% of samples. There is also an increased risk of genotyping errors due to higher salt concentrations or inhibitors. The melt curve analysis is highly sensitive to the salt concentration and variation of the salt concentration shifts the peak of the genotype relative to the controls.

Our DNA extraction from tissue uses spin columns from Qiagen to prepare the DNA. This guarantees the highest possible quality DNA for genotyping and downstream applications. Please send us between 3–5 mm of tail tissue or alternatively at least 3 mm² of earclip tissue.

Garvan's Genetics Core Facility was established in 2004 through support from the Australian Cancer Research Foundation (ACRF).

We offer the following services

Rodent genotyping, including establishing new mouse colonies
Copy number analysis (Het/Hom differentiation)
SNP analysis
Pathogen Testing.

Establishing new mouse genotyping projects

We will establish your current mouse genotyping PCR protocol on our system for $120. We can waive this fee depending on expected sample numbers that will be sent in future. Please fill in the New Project Submission Form. Please tell us who to report to, the gene name, the primer sequences, the primer combinations and product sizes and expected genotypes (e.g. wildtype, heterozygous, homozygous). We will also need some controls (HET, WT, HOM) to get us started. These controls can be DNA samples with known genotypes (confirmed by previous genotyping) or tissue from animals with known genotypes (e.g. founder animals).

We will order the primers and will establish the project once primers and controls have arrived. In case you have no primer sequences and know only the gene name or gene sequence, we can also design primers for your genotyping. We will be in contact with you along the way and inform you about the progress.

If your project is urgent and you need us to establish it quickly, you can provide us with primer aliquots and we will start genotyping immediately for you.

While it is the responsibility of each client to submit the correct primers (or sequence file to design primers) when establishing a new project, we can assist the process by making available our database of all the existing primers and PCRs that are currently established at GMG. Upon request, we can send you the document GMG Mouse Genotyping Existing PCRs and Primers, which contains a list of primers and PCRs and mouse lines that are already established at GMG.

Copy number analysis (Het/Hom differentiation)

When exactly the same amount of DNA is added into a PCR reaction and the amplification process monitored via realtime PCR, a homozygous sample will have a crossing point of one cycle earlier than a heterozygous sample. Therefore, we can distinguish heterozygous samples from homozygous samples for any given gene by this protocol. For the method to work we quantify the DNA exactly, dilute to the same concentration and set up five PCRs (repetitions) of each sample and two house keeper genes, amplify in realtime mode and form the average Ct value for each sample, subtract the house keeper and compare to controls. This test only works with very clean DNA and a het and hom control.

SNP analysis

We have developed a reliable and very competitively priced method to type SNPs in a high-throughput format via high-resolution melt curve (HRM) analysis.

Please mail the samples to Garvan Molecular Genetics via Australia Post at the address below.

Pathogen testing

We can test for several mouse pathogens. Currently established are:

Helicobacter ssp. (bilis, muridarum, hepaticus, typhlonius, rodentium)

Pasteurella ssp (Javetz, Heyl)
Corynebacterium bovis

We extract DNA from swabs and faecal samples.

Our services are provided through our sample submission portal. Visit the portal to create a new account, log into an existing account or find out more about submitting samples.

Before you submit a sample, please read the Mouse Genotyping Service Sample Submission Guidelines and Mouse Genotyping Service Questions & Answers. Our terms and conditions are available here.

For routine genotyping, we prefer samples to be in a 96 well format as these plates fit into our robotic liquid handlers. We can send you special plates for tissue or DNA samples. Please request this by emailing us.

Additional positive controls

Some researchers prefer to add positive controls to each plate they send us and for each test they want us to perform. We greatly appreciate this as it adds additional safety to the tests. If you can, please add positive and negative controls to your sample submission plate(s).

Personal sample delivery

How to submit your samples is described in detail in our Mouse Genotyping Service Sample Submission Guidelines. If you submit DNA samples the volume of the DNA sample should be at least 30 ul in each well of a 96 well plate with a concentration of at 5 ng/ul for standard genotyping. For copy number analysis we need at least 100 ul with 50 ng/ul concentration of the submitted DNA. We will not use all of the submitted DNA, we will most likely only use 1 ul of this DNA but we need this volume for the robotic liquid handlers to safely operate. If you submit tissue samples, please make sure they are between 3–5 mm of tail tissue or alternatively at least 3 mm² of earclip tissue.

Sample submission via Australia Post

You can also send your samples via Australian Post or courier to:

Garvan Molecular Genetics
Garvan Institute of Medical Research
Level 8

384 Victoria Street
Darlinghurst NSW 2010

Please email a Mouse Genotyping Sample Submission form to us with the billing information and an email address to which the results should be forwarded.

How we genotype your sample

We perform a touchdown realtime PCR in a 384 well plate scale. Samples are processed via liquid handlers and analysed on our LightCycler 480 with a Sito9-based melt curve analysis. The control melt curve peaks are compared to the sample peaks. We run positive and negative controls for each test and only release results when all controls have been validated.

Getting your results

The results of the genotyping will be sent to you via email. We usually complete the genotyping within 72 hours after receipt of sample. In the event of dropouts, we will repeat them once. If the genotype can still not be determined, we will inform you about the problem. In exceptional cases, the genotyping might be delayed by a day if instruments are out of order or staff is sick.

You can also label your samples as urgent and we will make a special effort to process them ASAP. Samples can be analysed with highest priority within 30 hours after receipt of sample, for an additional service fee.