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Quantitative Imaging of CRISPR-Labeled Genomic Loci, with the laboratory of David Grunwald, RNA Therapeutics Institute, UMass Medical School

This collaboration relates to imaging aspects of the first entry on the home page.

The Human Genome Repeated Sequence Database as Mined for CRISPR Labeling, with the laboratory of Shaojie Zhang, University of Central Florida

To obtain suitable markers for barcoding interphase chromosomes by our CRISPR labeling platforms we have collaborated with Dr. Zhang’s group to screen the entire repeated sequence component of the human genome for repeats that are 1: unique to a specific chromosome; 2: are present at some loci at sufficient copy number for detection by our labeling methods; and 3: are arrayed through the long  and short arms of the chromosomes in such a way as to allow us to relate our observed in vivo conformation of each chromosome with its DNA’s physical map.

Nucleolus-Associated Domains (NADs) in Murine Embryonic Stem Cells, with the laboratory of Paul Kaufman, Department of Molecular, Cell and Cancer Biology, UMass Medical School

Our laboratory is providing CRISPR-based labeling technology of NADs to complement Hi-C NAD mapping in the Kaufman lab.  The goal of this project is to examine how the nucleolus-adjacent regions of the genome changes as mES cells undergo differentiation along various pathways.  This project is part of the NIH 4D Nucleome Initiative.

DNA Single- vs. Double-Strand Break Repair Dynamics Imaged in Live Cells, with the laboratory of Jurek Drobrucki, Jagellonian University, Krakow, Poland

Using CRISPR Cas9 machinery that induces either a single- or double-strand break at a specific desired site, detected simultaneous by a fluorescent reporter, we are investigating the dynamics of repair factors at the site- each labeled in additional colors.  We believe we have an unprecedented opportunity to track these critical genome-protective events in a live cell context, to complement elegant biochemistry that has been done.

Role of Transcription-Coupled DNA Repair Proteins in the Puzzling Resistance of Cockayne Syndrome Patients to Skin Cancer, in collaboration with James Cleaver, University of California, San Francisco

Transcription-coupled DNA repair is a process in which specific proteins confer RNA polymerase II with the ability to transcribe over a gapped template DNA strand.  Patients with the condition Cockayne Syndrome are deficient in these proteins and also display a relative resistance to skin cancer.  The Cleaver laboratory has undertaken an systematic investigation of this question and we are providing cell lines in which each of the transcription-coupled DNA repair proteins has been edited out by CRISPR.

A Rapid Detection Method for c9orf72 Repeat Expansions as a New Diagnostic Tool in ALS, in collaboration with Zuoshang Xu, Department of Biochemistry and Molecular Pharmacology, UMass Medical School

In a major inherited form of the neurodegenerative disease amyotrophic lateral sclerosis (ALS) is a sequence in the c9orf gene undergoes a pathogenic expansion, which we have been able to detect in patient-derived cells by a novel, CRISPR-based cytological method that is simple to execute and takes less than one hour.  We are now refining this method and adapting it to cheek bucosal cell samples taken from subjects in which this form of ALS is suspected from the initial neurology clinic assessment.

Imaging Genome Activity in Barcoded Chromosomes at Nanoscale Spatial Resolution, in collaboration with the laboratory of Xiaowei Zhuang, HHMI and Department of Chemistry and Chemical Biology, Harvard University

A combination of MER-FISH (Zuang lab) and our CRISPRRainbow system is being deployed to track genome dynamics in a paired technology platform.

The Role of the Nuclear Lamina in Maintaining the Homeostasis of Nuclear Bodies, in collaboration with the laboratories of Sui Huang and Robert Goldman, Northwestern University Feinberg School of Medicine

We are testing the hypothesis of a functional connectome between the nuclear lamina and various nucleoplasmic structures including the nucleolus, Cajal Bodies and PML bodies.