A Common Human β Globin Splicing Mutation Modeled in Mice

Abstract

The βIVS-2-654 C→T mutation accounts for approximately 20% of β thalassemia mutations in southern China; it causes aberrant RNA splicing and leads to β0 thalassemia. To provide an animal model for testing therapies for correcting splicing defects, we have used the “plug and socket” method of gene targeting in murine embryonic stem cells to replace the two (cis) murine adult β globin genes with a single copy of the human βIVS-2-654 gene. No homozygous mice survive postnatally. Heterozygous mice carrying this mutant gene produce reduced amounts of the mouse β globin chains and no human β globin, and have a moderate form of β thalassemia. The heterozygotes show the same aberrant splicing as their human counterparts and provide an animal model for testing therapies to correct splicing defects at either the RNA or DNA level.

Use of mouse models to study the mechanisms and consequences of RBC clearance

“…Nonetheless, these SCD models have been instrumental in improving our understanding of vascular [105] and pulmonary complications [106] and for evaluating potential treatments [107]. Similarly, multiple mouse thalassaemia models, with various human mutations, are available [108][109][110][111][112][113][114][115][116].…”

Nanoparticle‐mediated genome editing in single‐cell embryos via peptide nucleic acids

“…To allow for analysis of gene editing both at the genotype and phenotype level, we used embryos derived from a transgenic reporter mouse (designated 654‐eGFP) containing a GFP‐beta globin fusion transgene with a IVS2‐654 (C to T) mutation in the beta globin‐derived intron causing an aberrantly spliced mRNA; correction by gene editing results in expression of a functional GFP mRNA transcript 22 . (Figure S2).…”

“…To allow for analysis of gene editing both at the genotype and phenotype level, we used embryos derived from a transgenic reporter mouse (designated 654‐eGFP) containing a GFP‐beta globin fusion transgene with a IVS2‐654 (C to T) mutation in the beta globin‐derived intron causing an aberrantly spliced mRNA; correction by gene editing results in expression of a functional GFP mRNA transcript. 22 (Figure S 2 ). To analyze levels of editing achieved in vitro, single‐cell fertilized embryos were harvested from homozygous 654‐eGFP mice and PLGA NPs containing PNA and donor DNA designed to correct the IVS2‐654 mutation 2 were pipetted into medium.…”

“…The 654‐eGFP mice used in the study were homozygous with two copies of a GFP‐beta globin fusion transgene with a IVS2‐654 (C to T) mutation causing an aberrantly expressed intron. Successful gene editing results in expression of a functional GFP mRNA transcript and eGFP fluorescence 22 …”

BACH1 regulates erythrophagocytosis and iron‐recycling in β‐thalassemia

“…WT (β m / m ) and β IVSII‐654 ‐thalassemic C57Bl/6 background mice (Lewis et al, 1998) were kindly supplied by the Thalassemia Research Center, Institute of Molecular Biosciences, Mahidol University, Salaya Campus, Thailand. This study was approved by the Ethics Review Board of the Institutional Animal Care and Use Committee of the Prince of Songkla University (approval number 2562‐10‐046).…”