SapTrap Assembly of Caenorhabditis elegans MosSCI Transgene Vectors.

Xintao Fan, Sasha De Henau, Julia Feinstein, Stephanie I Miller, Bingjie Han, Christian Frøkjær-Jensen, Erik E Griffin

Research output: Contribution to journalArticlepeer-review

2 Scopus citations

Abstract

The Mos1-mediated Single-Copy Insertion (MosSCI) method is widely used to establish stable Caenorhabditis elegans transgenic strains. Cloning MosSCI targeting plasmids can be cumbersome because it requires assembling multiple genetic elements including a promoter, a 3'UTR and gene fragments. Recently, Schwartz and Jorgensen developed the SapTrap method for the one-step assembly of plasmids containing components of the CRISPR/Cas9 system for C. elegans Here, we report on the adaptation of the SapTrap method for the efficient and modular assembly of a promoter, 3'UTR and either 2 or 3 gene fragments in a MosSCI targeting vector in a single reaction. We generated a toolkit that includes several fluorescent tags, components of the ePDZ/LOV optogenetic system and regulatory elements that control gene expression in the C. elegans germline. As a proof of principle, we generated a collection of strains that fluorescently label the endoplasmic reticulum and mitochondria in the hermaphrodite germline and that enable the light-stimulated recruitment of mitochondria to centrosomes in the one-cell worm embryo. The method described here offers a flexible and efficient method for assembly of custom MosSCI targeting vectors.
Original languageEnglish (US)
Pages (from-to)g3.400822.2019
JournalG3 (Bethesda, Md.)
Volume10
Issue number2
DOIs
StatePublished - Dec 17 2019

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