PhotoMorphs: a novel light-activated reagent for controlling gene expression in zebrafish.
Manipulating gene expression in zebrafish is critical for exploiting the full potential of this vertebrate model organism. Morpholino oligos are the most commonly used antisense technology for knocking down gene expression. However, morpholinos suffer from a lack of control over the timing and location of knockdown. In this report, we describe a novel light- activatable knockdown reagent called PhotoMorph. PhotoMorphs can be generated from existing morpholinos by hybridization with a complementary caging strand containing a photocleavable linkage. The caging strand neutralizes the morpholino activity until irradiation of the PhotoMorph with UV light releases the morpholino. We generated PhotoMorphs to target genes encoding enhanced green fluorescent protein, No tail, and E-cadherin to illustrate the utility of this approach. Temporal control of gene expression with PhotoMorphs permitted us to circumvent the early lethal phenotype of E-cadherin knockdown. A splice-blocking PhotoMorph directed to the rheb gene showed light-dependent gene knockdown up to 72 hpf. PhotoMorphs thus offer a new class of laboratory reagents suitable for the spatiotemporal control of gene expression in the zebrafish.
Tomasini
Amber J
AJ
Department of Pediatrics, Gastroenterology Section, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA.
Schuler
Aaron D
AD
Zebala
John A
JA
Mayer
Alan N
AN
D000818:Animals; D001483:Base Sequence; D015820:Cadherins / Q000235:genetics; D017931:DNA Primers; D005786:Gene Expression Regulation / Q000187:drug effects*; D055785:Gene Knockdown Techniques; D049452:Green Fluorescent Proteins / Q000235:genetics; D007202:Indicators and Reagents / Q000494:pharmacology*; D008027:Light; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015027:Zebrafish / Q000196:embryology / Q000235:genetics*
D015820:Cadherins; D017931:DNA Primers; D007202:Indicators and Reagents; D049452:Green Fluorescent Proteins