Human Genetic Disease and Genetic Engineering

XIST transgenes can silence an autosome into which they are integrated

chrm4The size of the full length genomic XIST transgene integrated into chromosome 4 (above) is ~35kb. However the full length 15kb cDNA works as well and we are currently testing the ability of a smaller 6.8kb transgene to silence.

 

Zinc Finger Nucleases (ZFN) provide highly efficient targeted transgene integration

Chrm19-XistPanel-ideogramUsing ZFN targeting technology we can aim XIST transgenes into different genomic environments and test the silencing capability of different regions (sequence and epigenetic content differences).

iPSC (induced pluripotent stem cells) provide a powerful alternative to human embryonic stem cells (hESC)

AP-DN25-4mouse-iPS mouseiPSwith-oct4

We are making both mouse and human iPS cells to study epigenetic changes in reprogramming and chromosome inactivation. The cells above are colonies of iPSC which we have made from mouse models of human disease.

Translating dosage compensation to trisomy 21

Trisomy 21 SilencingConcept for translating dosage compensation to trisomy 21: using genome editing with zinc finger nucleases, we targeted a large, inducible XIST transgene into the one of three Chromosome 21s in induced pluripotent stem cells (iPSCs), derived from Down syndrome patient cells. XIST RNA coats the extra chromosome 21 and triggers chromosome-wide transcriptional silencing and stable heterochromatin modifications, to form a “Chromosome 21 Barr Body.” 

Nature, July 2013, published online