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Gene Targeting

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Gene cloning (in 2 months, or less)
- construct the probe, screen the library and isolate clones (genomic BAC or cDNA), or retrieve appropriate sequences from public libraries
- verify clone inserts by Southern and/or PCR technology and appropriate sequencing
Desired origin (mouse strain):
129/Sv C57BL/6N BALB/c
Other (specify):
Is the full sequence known/published:
Yes Don’t know No
Type of sequence:
cDNA Genomic
Remarks:

Vector construction (in 5-6 months, or less)
- establish the targeting and PCR screening strategies, subclone DNA fragments
- construct the vector (& PCR control vector for targeting in embryonic stem cells)

For complex vectors, please contact either:
stefan.selbert@polygene.ch Tel. (direct line) +41 44 828 63 83 or
doron.shmerling@polygene.ch Tel. (direct line) +41 44 828 63 82

Vector type:
Conventional KNOCK-OUT:
Conditional KNOCK-OUT:
Size of the region to flank with lox sites: kbp
Knock-In (Insertion vectors)
Marker(s) to insert (e.g. Neo, GFP, …):
Expression Vectors
Promoter, transgene (cDNA) and regulatory elements (e.g. NLS, introns or poly-A)
all sequences available some fragments to be isolated by PolyGene
include inducible transgenic strategies
Screening strategy and PCR analysis by:
PolyGene Yourself
Remarks:


ES cell transfection (in 2-3 months, or less)
- electroporation of the targeting construct in ES cells and selection for antibiotic resistance,
- isolation & screening of ES cell clones, karyotyping & morphological clone description.

... optionally (within 2 months), further recombination for conditional transgenesis includes

- Electroporation of Cre or Flp for transient expression in ES cells (e.g. excision of selection markers),
- isolation & screening of ES cell clones, karyotyping & morphological clone description
Desired ES cell strain:
129/Sv C57BL/6N BALB/c
Other (specify):

Type of transgenesis:
Conventional KO Conditional KO Knock-In

Screening strategy and PCR analysis by:
Polygene Yourself

Type of screening strategy foreseen:
Southern: PCR
Remarks:

Blastocyst injection (in 2 months or less)
- isolation of blastocysts from appropriate donor mice, ES cell clone injection into blastocysts
- transfer of blastocysts (> 50) into fosters, and progeny coat colour determination
ES cell strain:
129/Sv C57BL/6N BALB/c
Other (specify):
Passage number:
Karyotype:
Remarks:

Please also send me information on:

ES cell in-vitro-differentiation (in 3-6 months or less)
- Start from isolated ES cell clones, and differentiation into any of a variety of cell types, such as cardiomyocytes, adipocytes, chondrocytes, epithelial-, neuronal-, smooth muscle-, skeletal muscle-, and blood cells, ...
- Analysis of marker/transgene expression (Western, Southern, RT-PCR or immunofluorescence)
Adipocytes Cardiomyocytes Chondrocytes
Epithelial Neuronal Skeleto-muscular
Blood Other (specify):

Required analysis:
Western Immunofluorescence
RT-PCR Northern
Other (specify):
Remarks:










PolyGene AG
Riedmattstrasse 9
CH-8153 Rümlang
Phone: +41 44 828 63 80
Fax: +41 44 828 63 81