Animal Models Service

We are specialized in generating transgenic mice and rats with the traditional method of pronuclear microinjection, as well as using lentiviral vectors. Animals generated in our facility possess SPF (Specific Pathogen Free) health status. Additionally, we offer the cryopreservation, embriotransfer, IVF and genotyping service.

Genotyping

The Genetic Engineering Core Facility offers genotyping service. Genomic DNA is extracted from the tissue fragment using comercially avaiable Genomic Mini Kit (A&A Biotechnology) and PCR (Polymerase Chain Reaction) is used for mutation detection using Qiagen or Syngen Taq Polymerase Kit. The genotyping protocol is avaiable here.

For prices and additional information please contact us:

ma.lewandowska@nencki.edu.pl

k.matusik@nencki.gov.pl

In vitro fertilization

In vitro fertilization is a method used for recovery of mice line from cryopreserved sperm . It may also be used as a method to overcome male fertility problems.

In vitro fertilization can be used to produce large colonies of mice in a relatively short period of time, saving also on mouse caging space. However, the success of IVF relies largely on that reasonable number of motile sperm can be retrieved. Therefore considering this method one should be awared, that reproductive differences between mouse strains seen in vivo are also evident in vitro. Males used for cryopreservation and IVF should be at age between 3-5 months and should have already proven fertility before.

The costumer provides at least 3 probes, each containing at least 10 ul of frozen sperm. The core will isolate oocytes from at least 10 superovulated females, perform IVF and transfer the resulting embryos to recipient females.

Do not hesitate to contact us for any questions:

j.przybys@nencki.edu.pl

Embryotransfer

Rederivation is a service of converting valuable transgenic lines into Specific Pathogen Free (SPF) status.

The embryos from animals, which are carrying or suspected of carrying pathogenic agents are removed and transferred into pathogen free animals. The embryo transfer method is the only one, which guarantees the removal of adventitious agents (viral, bacterial and parasite contamination) from your colony giving you pathogen free animals to work with.

We offer 2 options to rederive the line:

Option 1 - Heterozygotes rederivation (standard procedure) - Investivagor provides 3-5 transgenic males (2-6 month old, sexually mature and preferable previously successfully bred males). The Facility provides 5-20 donor females (6-8 weeks old) (C57Bl6/J, FVB, Wistar).

Option 2 - Homozygotes rederivation (recommendated for specific genetic background) - Investivagor provides 3-5 transgenic males as described above and ~10-20 donor females (6-8 weeks old).

After hormone-induced superovulation and mating, embryos are collected and transferred into foster mothers (mice CD1, rats SD). The tissue fragment from offspring will be collected for genotyping.

Clients must supply the following:

Completed rederivation protocol (Embryo Transfer.pdf )
Animals: at least 3 males of the strain to be rederived for standard procedure and both males and donor females for homozygote rederivation (or specific genetic background)
For rederivation from the frozen embryos the Investigator provides at least 100 frozen embryos (Line Reconstitution from Cropreserved Embryos.pdf)

Special notes

The standard price of the embryo transfer service includes up to 3 attempts of the line rederivation.

If the provided males fail to mate after 2 attempts or there will be no fertilized embryos, the investigator will be notified.

If the foster females fail to deliver live pups, the process will be repeated at no additional cost (other than the cost of more female egg donors, if needed). The investigator must inform the facility if the strain exhibits any development problems, severe defects or lethality, so that the pups can be properly monitored.

If the foster females fail to deliver pups of the proper genotype, the process will be repeated at no additional cost (other than the cost of more female egg donors, if needed) AFTER the investigator has done appropriate genetic testing on the 'dirty' animals to confirm their genotype.

If the investigator has not contacted the facility with the results of the genotype by weaning, all the pups will be transferred to the investigator's protocol number and account.

If the rederivation fails, the process will be repeated at no additional cost.

Submission form can be sent together with animals, as well as scanned and e-mailed in advance.

Do not hesitate to contact us for any questions:

j.przybys@nencki.edu.pl

a.klejman@nencki.edu.pl

Transgenic Animals Technology Services

The Transgenic Unit is a full-service core that provides transgenic and genome engineering services to enable the production of animal models by classical transgenesis and by viral vector modifications. We are performing both mice and rat transgenesis.

From 2017, supported by Foundation of Polish Science TEAM TECH CORE FACILITY PLUS grant - the unique integrated platform (ANIMOD) has been available for generation and analysis of animal models for biomedical applications.

Transgenesis of the animals is performed using the classical microinjection, improved in our laboratory (patent number 355 353). The first step is the preparation of the DNA. Plasmid is digested by restriction enzymes, to obtain a linear form. Then this linear form is microinjected to the prenuclei by the use of the micromanipulator. Then those hypothetically transgenic zygotes are implemented into tubal infundibulum of the pregnant females. After the birth the offspring is genotyped and if any transgene animal is found, it becomes the founder of a new, transgenic strain.

For more information and ordering details please contact us:

j.przybys@nencki.edu.pl

a.klejman@nencki.edu.pl

Please fill the transgenic animal generation order form.

Cryopreservation

Cryopreservation is cost and time saving method enabling preservation of valuable transgenic lines that are not currently in use.

Freezing of sperm or embryos is always recommended as they constitute as a ‘back-up’ in case of disease outbreak, fertility problems, spontaneous phenotype loss, etc. A complete cycle of cryopreserving a strain and reconstituting it from frozen material involves the same biological steps whether one uses embryos or sperm. In both cases, the production of fertilized embryos is the most expensive step. With embryo cryo, the fertilization step takes place before freezing, whereas with sperm cryo it is done after thawing.

There are 2 possible embryo cryopreservation options depending on mice genotype:

Option 1 - Cryopreservation of heterozygous animals. Investigator provides 5-10 stud males (3-6 months old). The Facility provides ~30 donor females (C57Bl6/J or FVB), which allowing to collect ~ 300 embryos/line at morula/blastocyst stage of development.

Option 2 - Cryopreservation of homozygous animals. Investigator provides 5-10 stud males (3-6 months old) and ~20 donor females (5-8 weeks old), enabling to receive ~200 morulas/blastocysts.

Embryo Freezing Order Form

For sperm cryopreservation we will freeze the sperm isolated from 3-5 practiced males (3-6 month of age) per line

Sperm Cryopreservation Order Form

For more information and ordering details please contact us:

j.przybys@nencki.edu.pl

a.klejman@nencki.edu.pl

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