An infected mouse or rat strain can be cleaned from pathogens/parasites by embryo transfer. This involves flushing preimplantation embryos (morula stage) and transferring them to a clean pseudopregnant recipient. Common viruses and bacteria infecting laboratory rodent strains include Mouse Hepatitis Virus (MHV), Parvovirus, Helicobacteria, Pasteurella.

Instructions for the customers:

You need for cleaning of a mouse strain 4-8 males and for a rat strain 2-4 males of the strain to be cleaned.

NOTE! The Males should be under 8 mo of age. If they are older, please contact the GM unit for instructions.

Schedule the pathogen removal with the GM mouse unit.

Fill in the work request form and send it to GM-unit. Please provide:

  • The name of the strain and the project license number or activity report number
  • Information on the location, identification and date of birth of the males to be used for pathogen removal
  • Genetic background of the females to be used (usually the same as the genetic background of the males). Note! GM unit will order the wild type females.

 

The GM unit will:

  • Schedule the pathogen removal and order wild type females needed.
  • Superovulate and mate the females and dissect oviducts for embryo collection
  • Wash the obtained embryos and transfer them to the clean foster mothers

Note! If the mouse/rat line to be cleaned is located in Biomedicum animal facilities, the Researcher has to deliver oviducts to GM-unit to F-building in Viikki according previously agreed schedule. Infected oviucts may not be brought to the F-building animal facilities, but they are handed to the personel outside the unit. Contact personnel for details.

The personnel of the animal facility will take the genotyping samples of the offspring resulting from the embryo transfer and send them to you.

After obtaining the genotyping results, inform both GM unit (gm-unit -at- helsinki.fi) and the animal facility personnel via Provet. After this the animal facility will take care of the shipment of health monitoring samples of the cleaned strain.

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A rodent strain can be archived as frozen preimplantation embryos (morulae). They can be stored in liquid nitrogen for at least 10 years. In this service ca. 250 embryos are frozen. This is enough for at least two strain retrievals. Successful freezing is controlled by a test retrieval from which the researcher genotypes the pups being born.

Shipment of genetically modified strains as frozen embryous is currently the best way to transport animalsy. This helps with problems such as pathogen infection, shipment of live animals etc.

About 250 mouse embryos can usually be frozen in 4-5 weeks while in rats this takes longer (some 6-8 weeks). If serious problems arise (no or few embryos recovered, no plugs, etc.), the process is stopped and the customer is informed and billed for the expenses.

Instructions for the customers

For mouse embryo freezing you need 6-8 males/strain and for rat 2-4 males/strain.

NOTE ! If the males are older than 8 mo, please contact the GM unit for instructions

Schedule the embryo freezing with the GM mouse unit.

Fill in the work request form and send it to GM-unit. Please provide:

  • The name and the project license number or activity report number of the strain to be frozen
  • Information on the location, identification and date of birth of the males (6-8) to be used for embryo freezing
  • Genetic background of the females to be used (Usually same as the genetic background of the males). Note! GM unit will order the females.

 

After freezing of embryos, some of them will be thawed and transferred to pseudopregnant females to control the success of the freezing. Genotyping samples from the resulting offspring will be sent to the customer. Please send the genotyping results to the GM unit (gm-unit -at- helsinki.fi). Note! Only after genotyping confirmation the males used for cryopreservation can be sacrificed.

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As mouse strain can also be cryopreserved as frozen sperm collected from fertile (homo- or heterozygous) males.

To order the service, please fill in a request form and send it to the GM unit.

 

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Genitically modified embryonic stem cells, ES cells, can be aggregated with morula-stage preimplantation embryos to generate chimeric mice transmitting the targeted mutations through the germ line. In addition to the locally produced ES-cells, cells with targeted or "gene trap" alleles produced by the international mutagenesis consortia can be aggregated (including ES cells in C57BL/6N background).

The morula-aggregation service provides germ line transmission or about 120 transferred aggregates/ES cell line.

Instructions for the customer

For morula-aggregation, ES cells are needed.

GM unit cultures ES cells for morula-aggregation in F-building in Viikki. Is so wanted, the researcher can do the ES cell culture work him/herself and provide the cells to th GM unit. See Culture protocol.

Primary embyonic fibroblast cells (both wt and neo resistant) for culturing the ES cells can be purchased from the GM unit.

Schedule morula aggregation with the GM unit

Fill in the work request form and send it to GM-unit. Please provide:

  • The name and the project license number of the strain to be generated
  • Information on the ES cell line (Name of the parental line and the genetic background, for example (R1-129sv)).

Note! If the researcher cultures the the ES cells, it has to be noted that the GM mouse unit will trypsinize the cells and make dilutions for aggregations.

When the chimeras are weaned, the sex, number and ES cell contribution (% of chimerism) will be determined. The customer will inform the animal facility for the subsequent breeding of the chimeras.

 

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The clustered, regularly interspaced, short palindromic repeats (CRISPR)-associated protein (Cas) technology has a revolutionized genome editing especially in animal transgenesis. The system consists of the Cas9 nuclease and a guide RNA, which guides the Cas9 enzyme to the target sequence to introduce double stranded breaks in genomic DNA. CRISPR/Cas9 technology enables mouse genome manipulation with exceptional simplicity, speed and superior efficiency.

Genomic manipulation with CRISPR/Cas9 technology utilizes either pronucleus or cytoplasmic injections of reagents. Our facility uses both of these techniques, but our routine work is based on cytoplasmic injections with commercial Cas9 as mRNA or protein included in the service.

Schedule CRISPR/Cas9 injection with the GM unit

Fill in the work request form and send it to GM-unit. Please provide:

  • The name and the project license number of the strain to be generated
  • Information on the mouse / rat strain to be used as oocyte donors (FVB, C57BL/6 or Wistar, Sprague-Dawley).

 

Take the prepared guideRNA(s) (and potential DNA construct) to the GM unit as scheduled

The animal facility personnel will take the genotyping samples from putative founder animals born and send them to you. Please send the genotyping results both to the GM unit and the animal facility, and inform the animal facility of the continuation.

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DNA pronuclear injection

Transgenic rodents are produced by pronuclear injection technique, in which DNA solution is injected in one of the pronuclei of a fertilized oocyte.

In the DNA pronuclear injection service approximately 250 oocytes are injected and transferred to a pseudopregnant female.

Instructions for the customers

For the DNA pronuclear injection you need linearized DNA construct. Protocol

Schedule DNA pronuclear injection with the GM mouse unit

Fill in the work request form and send it to GM-unit. Please provide:

  • The name and the project license number of the strain to be generated
  • Information on the mouse/rat strain to be used as oocyte donors (FVB, C57BL/6 or Wistar, Sprague-Dawley). FVB is the most frequently used strain in mice.
     

 

Take the prepared DNA construct to the GM mouse unit as scheduled

The animal facility personnel will take the genotyping samples from putative founder animals born and send them to you. Please send the genotyping results both to the GM unit and the animal facility and inform the animal facility of the continuation.

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Mouse strains can be shipped as frozen preimplantation embryos, or fozen sperm. When shipping frozen material, provide the recipient with the thawing protocol.

The GM unit helps customers with shipment, but will not take care of the arrangements of the shipment.

When shipping frozen material, you need a special container (so called dry shipper), which you can get from the recipient or from the GM unit.

Speed Congenics method accelerates the change of the genetic background by backcrossing and reduces the amounts of mice needed.

The genotyping services are produced by the Molecular Medicine Sequencing Laboratory (see further information)

A mouse strain can be recovered by thawing cryopreserved sperm that is used to fertilize wild type oocytes (collected from superovulated females) in vitro. The resulting two-cell embryos are transferred to pseudopregnent recipient females.

To order the service, please fill in a request form and send it to the GM unit.

 

Pricelist

The GM unit provides custom services also in other procedures related to preimplantation embryos. The GM unit has experience on tetraploid aggregation, BAC DNA and sytoplastic injection, production of new ES cell lines, etc. Billing is according to time and material spent for the project.