In vivo expression of mutant calpains in the eye using lentivirus

Katherine J. Wert, Vinit B. Mahajan

Research output: Chapter in Book/Report/Conference proceedingChapter

1 Scopus citations

Abstract

Exome sequencing has identified many candidate genes and mutations for human diseases, but the functional validation of these candidates is a time-consuming and costly process. Here, we describe a method which uses lentiviruses to overexpress calpain mutations that may play a role in dominant diseases such as autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV). The use of lentivirus to deliver the mutant calpain allows for a cost-effective, rapid, and efficient approach to test whether or not a candidate gene mutation from exome sequencing acts as the disease-causing allele for a human disorder. This method also provides for a comparison of different candidate mutations from a single gene identified by exome sequencing, as well as elucidating the mechanisms underlying these complex human disorders. Furthermore, this chapter focuses on two different methods to deliver mutant calpain to the cells of the eye, using either a subretinal or an intravitreal injection of the lentivirus into the mouse eye.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages233-247
Number of pages15
DOIs
StatePublished - Jan 1 2019
Externally publishedYes

Publication series

NameMethods in Molecular Biology
Volume1915
ISSN (Print)1064-3745

Keywords

  • Calpain
  • CAPN5
  • Intravitreal injection
  • Lentiviral vectors
  • Molecular cloning
  • Mouse eye
  • Subretinal injection

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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  • Cite this

    Wert, K. J., & Mahajan, V. B. (2019). In vivo expression of mutant calpains in the eye using lentivirus. In Methods in Molecular Biology (pp. 233-247). (Methods in Molecular Biology; Vol. 1915). Humana Press Inc.. https://doi.org/10.1007/978-1-4939-8988-1_18