ES Cell Targeting

Responsibilities of the Investigator

  • Discuss experimental design with facility directors before beginning your project.
  • Provide a map of the construct including size, regions of homology, selection strategy, enzyme sites and oligo binding sites.
  • Identify a unique restriction site to linearize your plasmid.  Provide gel documentation of your test digest.
  • Provide 100ug of purified plasmid at 1ug/ul.
  • Develop a reliable genotyping assay (long PCR, Southern, or both) before submission of DNA to the facility.
  • Screen picked clones for targeted candidates within three months (during which time the clones are stored at -80o).
  • Confirm genotype of candidate clones by Southern blot.
  • Submit a Request Form ESC to initiate the project.

Services Provided by the Facility

  • The gene-targeting facility will linearize your plasmid, electroporate into ES cells, perform selection and pick clones.
  • ES cell lines provided:
    • E14 (129/Ola parental ESC line)
    • EDJ22 (129/SvEv parental ESC line)
    • JM8.N4 (C57Bl/6N parental ESC line)
    • JM8.A3 (C57Bl/6N agouti-rescued parental ESC line)
  • We guarantee two rounds of electroporations (if necessary) for each targeting construct.
  • Picked clones will be stored at -80o for up to three months, during which time the investigator must screen for candidate clones.
  • The facility will expand candidate clones for freezing and confirmation of genotyping by Southern blot (investigator provides Southern blot confirmation).
  • After confirmation of homologous recombination, the facility will karyotype up to four of these clones.  Any additional expansion and/or karyotyping of clones will be subject to an additional charge.

Fee for ES Cell Targeting Service

  • $4500.00 ES cell targeting (includes expansion and karyotyping of screened ES cells).

REQUEST FORM ESC

Last modified: May 14, 2014
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