These mice harbor the hα (Hba^tm1(HBA)Tow) targeted mutation, the -1400 γ-β^S (Hbb^{tm2(HBG1,HBB*)Tow}) targeted mutation, and/or the -383 γ-β^A (Hbb^{tm3(HBG1,HBB)Tow}) targeted mutation. These mice should not harbor any wildtype allele at the Hbb locus.

The Hba^tm1(HBA)Tow targeted mutation (hα) was created by replacing the endogenous mouse α-globin gene with a human α-globin gene.

The Hbb^{tm2(HBG1,HBB*)Tow} targeted mutation (-1400 γ-β^S) was created by replacing the endogenous mouse major and minor β-globin genes with both the human ^Aγ gene (with 1400 bp of 5' flanking sequence) and the human β^S-globin gene (-1400 γ-β^S). The β^S gene contains an A to T mutation in the sixth codon of the human β-globin gene (resulting in a glutamic acid to valine conversion) that has been linked to sickle cell disease.

The Hbb^{tm3(HBG1,HBB)Tow} targeted mutation (-383 γ-β^A) was created by designing a targeting construct with the human hemoglobin gamma (^Aγ) gene with 383 bp of 5' flanking sequence, the human wildtype hemoglobin beta (β^A) gene with 815 bp of 5' flanking sequence, and a loxP-flanked phosphoglycerase/Hygromycin (PGK/Hygro) selection cassette.

The -383 γ-β^A targeting construct was electroporated into embryonic stem (ES) cells from knock-in sickle cell mice (homozygous for the hα targeted mutation and homozygous for the -1400 γ-β^S targeted mutation). The -383 γ-β^A construct contains 383 bp of 5' flanking sequence which is targeted to 1400 bp of 5' flanking sequence already present in the ES cells as part of the -1400 γ- β^S allele. Different recombination events in this flanking region produced two ES cell clones; clone 2 harbored the -383 γ-β^A replacement allele. This clone was then transiently transfected with a Cre expressing plasmid to delete the PGK/Hygro cassette, and then injected into recipient blastocysts. The resulting chimeric males were bred to females homozygous for hα and heterozygous for -1400 γ-β^S. Offspring with the sickle cell corrected genotypes (homozygous for hα and heterozygous at the Hbb locus (-383 γ-β^A/-1400 γ-β^S)) were obtained. These mice were maintained on a mixed C57BL/6;129 genetic background by breeding together for many generations prior to sending to The Jackson Laboratory Repository as Stock No. 013071.

Dr Paul Schmidt, from Boston Children's Hospital, purchased some Stock No. 013071 mice and subsequently separated the three alleles in order to backcross the strain to C57BL/6J (Stock No. 000664). Each allele was bred to C57BL/6J for at least 10 generations before being bred back together in one strain. The C57BL/6J-congenic strain was sent to The Jackson Laboratory as Stock No. 027265.

• Homozygous for Hba^tm1(HBA)Tow, Homozygous for Hbb^{tm3(HBG1,HBB)Tow} (non-sickling from the colony)

Approximate Controls

• 000664 C57BL/6J

Additional Information

• Considerations for Choosing Controls

• Wu LC; Sun CW; Ryan TM; Pawlik KM; Ren J; Townes TM. 2006. Correction of sickle cell disease by homologous recombination in embryonic stem cells. Blood 108(4):1183-8PubMed: 16638928MGI: J:134980