Identification of zebrafish shield organizer-specific genes
10.3969/j.issn.1005-4847.2016.05.001
- VernacularTitle:斑马鱼胚胎胚盾特异表达基因的鉴定
- Author:
Chuanlu WAN
;
Yifang YAN
;
Yu CAO
;
Qiang WANG
- Publication Type:Journal Article
- Keywords:
Zebrafish;
Shield organizer;
FACS;
RNA deep sequencing;
in situ hybridization
- From:
Acta Laboratorium Animalis Scientia Sinica
2016;24(5):441-447
- CountryChina
- Language:Chinese
-
Abstract:
Objective During zebrafish gastrulation, dramatic movements rearrange cells into three germ layers and contribute to the formation of the shield organizer, which acts as a dorsal signal center to pattern the body axis.Global identification of shield organizer-specific genes in early gastrulas will be valuable for studying the regulatory cascades during organizer formation and body axis establishment.Methods Tg( gsc:GFP) transgenic embryos express GFP in the shield organizer, which is controlled by a 1.8 kb gsc promoter.Flow cytometry technology and RNA deep sequencing analysis were applied to isolate GFP positive cells from the Tg( gsc:GFP) transgenic embryos and systematically uncover the genes highly expressed in the dorsal organizer.Subsequently, the expression of shield organizer-specific genes was further con-firmed by quantitative real-time PCR and whole mount in situ hybridization method during zebrafish embryonic develop-ment.Results GFP-positive cells exceeding 96%purity were isolated from shield-stage Tg(gsc:GFP) transgenic embryos and 657 organizer highly expressed genes were identified through RNA deep sequencing analysis.The results of in situ hy-bridization experiments revealed that a number of genes including KIAA1324, ripply1, twist2, isthmin1, nme4, zgc174153 and rrbp1b were expressed in shield organizer during zebrafish gastrulation.Conclusions The identification of these shield organizer-specific genes in the current study provides useful clues to explore the zebrafish developmental functions in further studies.
