A medium-scale assay for enhancer validation in amniotes

TY - JOUR

T1 - A medium-scale assay for enhancer validation in amniotes

AU - Chen, Jingchen

AU - Streit, Andrea

N1 - © 2015 The Authors. Developmental Dynamics published by Wiley Periodicals, Inc. on behalf of American Association of Anatomists.

PY - 2015/7/15

Y1 - 2015/7/15

N2 - BACKGROUND: Enhancers are key elements to control gene expression in time and space and thus orchestrate gene function during development, homeostasis, and disease. Whole genome approaches and bioinformatic predictions have generated a tremendous pool of potential enhancers, however their spatiotemporal activity often remains to be validated in vivo. Despite recent progress in developing high throughput strategies for enhancer evaluation, these remain mainly restricted to invertebrates and in vitro cell culture.RESULTS: Here we design a medium-scale method to validate potential enhancers in an amniote embryo, the chick. Using a unique barcode for different reporter vectors allows us to detect the activity of nine separate enhancers in a single embryo by one-step RT-PCR. The assay is sufficiently sensitive to expand its capacity further by generating additional barcoded vectors.CONCLUSIONS: As a rapid, sensitive, and cost-effective way to assess enhancer activity in an amniote vertebrate, this method provides a major advance and a useful alternative to the generation of transgenic animals. Developmental Dynamics, 2015. 

AB - BACKGROUND: Enhancers are key elements to control gene expression in time and space and thus orchestrate gene function during development, homeostasis, and disease. Whole genome approaches and bioinformatic predictions have generated a tremendous pool of potential enhancers, however their spatiotemporal activity often remains to be validated in vivo. Despite recent progress in developing high throughput strategies for enhancer evaluation, these remain mainly restricted to invertebrates and in vitro cell culture.RESULTS: Here we design a medium-scale method to validate potential enhancers in an amniote embryo, the chick. Using a unique barcode for different reporter vectors allows us to detect the activity of nine separate enhancers in a single embryo by one-step RT-PCR. The assay is sufficiently sensitive to expand its capacity further by generating additional barcoded vectors.CONCLUSIONS: As a rapid, sensitive, and cost-effective way to assess enhancer activity in an amniote vertebrate, this method provides a major advance and a useful alternative to the generation of transgenic animals. Developmental Dynamics, 2015. 

U2 - 10.1002/dvdy.24306

DO - 10.1002/dvdy.24306

M3 - Article

C2 - 26177930

SN - 1058-8388

VL - 244

SP - 1291

EP - 1299

JO - Developmental dynamics : an official publication of the American Association of Anatomists

JF - Developmental dynamics : an official publication of the American Association of Anatomists

IS - 10

ER -