Institut for Biomedicin

Lars Bolund

CRISPR-C: circularization of genes and chromosome by CRISPR in human cells

Publikation: Bidrag til tidsskrift/Konferencebidrag i tidsskrift /Bidrag til avisTidsskriftartikelForskningpeer review

DOI

  • Henrik Devitt Møller, Department of Veterinary Disease Biology, University of Copenhagen, Frederiksberg, Denmark; Zoology Department, Faculty of Science, Cairo University, Giza, Egypt.
  • ,
  • Lin Lin
  • Xi Xiang
  • Trine Skov Petersen
  • Jinrong Huang, China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China.
  • ,
  • Luhan Yang, eGenesis, Inc., Cambridge, MA 02139, USA.
  • ,
  • Eigil Kjeldsen
  • Uffe Birk Jensen
  • Xiuqing Zhang, BGI Education Center, University of Chinese Academy of Sciences, Shenzhen 518083, China., BGI-Shenzhen, 518083 Shenzhen, China; China National GeneBank-Shenzhen, BGI-Shenzhen, 518083 Shenzhen, China., China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China.
  • ,
  • Xin Liu, China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China.
  • ,
  • Xun Xu, China National GeneBank, BGI-Shenzhen, Shenzhen 518120, China.
  • ,
  • Jian Wang, James D. Watson Institute of Genome Science, 310008 Hangzhou, China.
  • ,
  • Huanming Yang, James D. Watson Institute of Genome Science, 310008 Hangzhou, China.
  • ,
  • George M Church, Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
  • ,
  • Lars Bolund
  • Birgitte Regenberg, Department of Veterinary Disease Biology, University of Copenhagen, Frederiksberg, Denmark; Zoology Department, Faculty of Science, Cairo University, Giza, Egypt.
  • ,
  • Yonglun Luo

Extrachromosomal circular DNA (eccDNA) and ring chromosomes are genetic alterations found in humans with genetic disorders. However, there is a lack of genetic engineering tools to recapitulate and study the biogenesis of eccDNAs. Here, we created a dual-fluorescence biosensor cassette, which upon the delivery of pairs of CRISPR/Cas9 guide RNAs, CRISPR-C, allows us to study the biogenesis of a specific fluorophore expressing eccDNA in human cells. We show that CRISPR-C can generate functional eccDNA, using the novel eccDNA biosensor system. We further reveal that CRISPR-C also can generate eccDNAs from intergenic and genic loci in human embryonic kidney 293T cells and human mammary fibroblasts. EccDNAs mainly forms by end-joining mediated DNA-repair and we show that CRISPR-C is able to generate endogenous eccDNAs in sizes from a few hundred base pairs and ranging up to 207 kb. Even a 47.4 megabase-sized ring chromosome 18 can be created by CRISPR-C. Our study creates a new territory for CRISPR gene editing and highlights CRISPR-C as a useful tool for studying the cellular impact, persistence and function of eccDNAs.

OriginalsprogEngelsk
TidsskriftNucleic Acids Research
Vol/bind46
Nummer22
Sider (fra-til)e131
ISSN0305-1048
DOI
StatusUdgivet - 14 dec. 2018

Se relationer på Aarhus Universitet Citationsformater

ID: 138766318