Development of a transgenic mouse line carrying a functional human Beta-globin gene with the IVSI-6 thalassemia mutation for in vivo screening and validation of aberrant splicing correctors

Mouse models that carry mutations causing thalassemia represent a suitable tool to test in vivo new mutation-specific therapeutic approaches. We report the production and characterization of a transgenic mouse line (TG-beta-IVSI-6) carrying the IVSI-6 thalassemia point mutation within the human beta-globin gene. At present, no "in vivo" models of this mutation are available. We characterized these transgenic mice generated by microinjection, with respect to their transgenic insertion, by PCR, Quantitative Multiplex PCR of Short fluorescent Fragments (QMPSF) and Fluorescence In Situ Hybridization (FISH). The transcription pattern was analysed by quantitative RT-PCR and the synthesis of hemoglobins by western blotting and Capillary Electrophoresis. (CE). In the TG-beta-IVSI-6 mouse (a) the transgenic integration region was located in mouse chromosome 7; (b) the expected altered splicing isoforms were found and (c) the transgenic ??-globin tissue specific gene expression, leady to the production of a chimeric mu alpha-globin2/hu beta-globin2 hemoglobin, was demonstrated. In conclusion, the TG-beta-IVSI-6 mouse reproduces molecular features of IVSI-6 beta-thalassemia and might be used as an in vivo model to characterize the effects of antisense oligodeoxynucleotides (ODN) targeting the criptic sites responsible for the generation of aberrantly spliced beta-globin RNA sequences, caused by the IVSI-6 mutation. These experiments are expected to be crucial for the development of a personalized therapy for beta-thalassemia.