Double UP: A Dual Color, Internally Controlled Platform for Knockdown or Overexpression

Overview

Journal Front Mol Neurosci

Specialty Molecular Biology

Date 2020 Jun 9

PMID 32508591

Citations 7

Authors

Russell J Taylor

Justin Carrington

Leah R Gerlach

Kendra L Taylor

Karl E Richters

Erik W Dent

Affiliations

Soon will be listed here.

Abstract

electroporation (IUE) is a powerful tool for testing the role of genes in neuronal migration and function, but this technique suffers from high degrees of variability. Such variability can result from inconsistent surgery, developmental gradients along both rostral-caudal and medial-lateral axes, differences within littermates and from one litter to another. Comparisons between control and experimental electroporations rely on section matching, which is inherently subjective. These sources of variability are cumulative, leading to difficult to interpret data and an increased risk of both false positives and false negatives. To address these limitations, we developed two tools: (1) a new plasmid, termed Double UP, which combines LoxP-flanked reporters and limiting Cre dosages to generate internal controls, and (2) an automated program for unbiased and precise quantification of migration. In concert, these tools allow for more rigorous and objective experiments, while decreasing the mice, time, and reagents required to complete studies.

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