University of Massachusetts

The United States of America, as Represented by the Secretary, Department of Health and Human Services

University of Central Florida Research Foundation Inc

Hanhui Ma

Thoru Pederson

Shaojie Zhang

Ardalan Naseri

Li-Chun Tu

Maximilaan Huisman

Scot Andrew Wolfe

Metewo Selase Kosi Enuameh

Nicola Anne Kearns

Rene Maehr

Manuel Garber

Ryan Michael Jude Genga

Tu Li-Chung

Chunxing Yang

DETECTION OF GENE LOCI WITH POLYCHROMATIC CRISPR-ASSOCIATED PROTEIN 9

The present disclosure relates to methods of and systems for modifying the transcriptional regulation of stem or progenitor cells to promote their differentiation or reprogramming of somatic cells. Further, the labeling and editing of human genomic loci in live cells with three orthogonal CRISPR/Cas9 components allow multicolor detection of genomic loci with high spatial resolution, which provides an avenue for barcoding elements of the human genome in the living state.

Compositions and methods for differential Cas9 gene labeling and/or editing

A C9orf72 DNA repeat expansion can be detected using a CRISPR Arrayed Repeat Detection System (CARDS). Based upon the compositions and methods supporting this platform primary cell cultures and/or blood cell smears can be tested under conventional clinical diagnostic laboratory conditions to diagnose genetically-based diseases having DNA repeat expansions, including but not limited to ALS. dCas9 constructs are also contemplated as having fluorescent proteins bound to any or all stem loop sequences, wherein detection of a plurality of dCas9 constructs having different colored fluorescent proteins can simultaneously detect at least six (6) different gene target loci.

Detection of gene loci with CRISPR arrayed repeats and/or polychromatic single guide ribonucleic acids

The present invention discloses a transcription activator-like effector-based strategy, termed 'TALEColor', for labeling specific repetitive DNA sequences in human chromosomes. TALEs were custom designed for human telomeric repeats and fused with any of numerous fluorescent proteins (FPs). TALE-telomere-FP fusion proteins were used to detect telomeric sequence in both living cells and fixed cells. Using human cells with different average telomere lengths, TALEColor signals correlated positively with telomere length. TALEs were also designed to detect centromeric sequences unique to specific chromosomes, enabling localization of these specific human chromosomes in live cells. These methods may have significant potential both for basic chromosome and genome research as well as in clinical applications.

Hybridization- independent labeling of repetitive DNA sequence in human chromosomes

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Shrewsbury, MA, United States of America

Hanhui Ma