Regulatory regions of the mouse Pax6 gene C a transcription Factor by itself Pancreas comea neural tube Enhancers C Exons 55a67 Map of enhancer sites of the Pax6 gene. The gene has 2 upstream promoters (in exon0 and exon 1)either of each can initiate transcription of the same RNA. Regions A-D are enhancer regions specific for different tissues B-glactosidase (reporter gene Transgenic mouse embryo An upstream enhancer(B)directs the acz reporter transgene in the surface ectoderm overlying the optic cup
An upstream enhancer (B) directs the LacZ reporter transgene in the surface ectoderm overlying the optic cup . Regulatory regions of the mouse Pax6 gene ( a Transcription Factor by itself ) Map of enhancer sites of the Pax6 gene . The gene has 2 upstream promoters ( in exon 0 and exon 1 ) either of each can initiate transcription of the same RNA . Regions A-D are enhancer regions specific for different tissues . Transgenic mouse embryo
TABLE 21.1 Some genes encoding human transcription factors and phenotypes resulting from their mutation Gene Mutation phenotype Androgen receptor Androgen insensitivity syndrome(Ch. 20) Azoosperm CBFAl Cleidocranial dysplasia(Ch. 15) CSX Heart defects EMX2 Schizencephaly(Ch. 23) Estrogen receptor Growth regulation problems, sterility( Ch 15) Forkhead-like 15 Thyroid agenesis, cleft palate GLⅠ3 Grieg syndrome( Ch. 16) HOXA-1 Hand-foot-genital syndrome(Ch. 16) HOXD-13 Polysyndactyly(Ch. 16) LMXIB Nail-patella syndrome( Ch 16 MITF Waardenburg syndrome type 2( Chs. 5, 21) PAX2 Renal-coloboma syndrome( Ch 15) PAX3 Waardenburg syndrome type 1( Ch 13) PAX6 Aniridia(Chs. 5, 21) PTX2 Reiger syndrome(Ch. 11) PITX3 Congenital cataracts POU3F4 Deafness and dystonia SOX9 Campomelic dysplasia, male sex reversal(Chs. 15, 20) SRY Male sex reversal(Ch 20 TBX3 Schinzel syndrome(uIna- mammary syndrome) TBX5 Holt-Oram syndrome( Ch. 16) TCOF Treacher-Collins syndrome TWIST Seathre-Chotzen syndrome WTI Urogenital anomalies( Ch 15)
Knock out mice In Knockout Mice it is essential, that the foreign dna is integrated into a specific dNa target within the mouse genome the gene which should be knocked out
Knock Out Mice In Knockout Mice it is essential, that the foreign DNA is integrated into a specific DNA target within the mouse genome = the gene which should be knocked out
The mouse Life span approx 2.5 years Gestation 21 days Litter size:8 to 12 Generation time three months Several inbred strains Genomic database Most advanced genetic technologies Over 90% identical to human genome Large enough for physiological studies
The MOUSE Life span: approx. 2.5 years Gestation : 21 days Litter size: 8 to 12 Generation time: three months Several inbred strains Genomic database Most advanced genetic technologies Over 90% identical to human genome Large enough for physiological studies
KNOCKOUT MOUSE VECTOR Gene of interest Exon 2 Exon 3 Exon 4 Eⅹon1 1000BP 5000BP( the green areas are homologous Knockout vector eo r HSVTk Neo r, a gene that encodes an HSVtk= Herpes Simplex Virus thymidine kinase: tk, a enzyme that inactivates the gene that encodes thymidine kinase, an enzyme that antibiotic neomycin and its phosphorylates the nucleoside analog ganciclovir (9 relatives, like the drug G418 (1, 3-dihydroxy-2-propoxy )methylguanine) which is lethal to mammalian cells DNa polymerase fails to discriminate against the (Protein synthesis, translation resulting nucleotide and inserts this nonfunctional nucleotide into freshly-replicating DNA (DNA replication is interrupted). So ganciclovir kills cells that contain the tk Final result( cells are resistant to Neomycin and Gancyclovir insensitive Neor Eⅹon1 Exon 3 Exon 4
KNOCKOUT MOUSE VECTOR . Exon 1 Exon 2 Exon 3 Exon 4 Neo r HSVtk Knockout vector HSVtk= Herpes Simplex Virus thymidine kinase : tk, a gene that encodes thymidine kinase, an enzyme that phosphorylates the nucleoside analog ganciclovir (9- (1,3-dihydroxy-2-propoxy)methylguanine) . DNA polymerase fails to discriminate against the resulting nucleotide and inserts this nonfunctional nucleotide into freshly-replicating DNA (DNA replication is interrupted ) . So ganciclovir kills cells that contain the tk gene. Neo r, a gene that encodes an enzyme that inactivates the antibiotic neomycin and its relatives, like the drug G418, which is lethal to mammalian cells; (Protein synthesis, translation) Final result ( cells are resistant to Neomycin and Gancyclovir insensitive ) Exon 1 Exon 3 Exon 4 Neo r 1000 BP 5000BP ( the green areas are homologous ) Gene of interest