title: NextSeq 500 sequencing; ﻿Anatomical and molecular dissection of the prestreak chick embryo
design: ﻿Anatomical and molecular dissection of the prestreak chick embryo
library_construction_protocol: Prestreak XII-XIII embryos (Eyal-Giladi and Kochav's stages described here DOI: 10.1016/0012-1606(76)90178-0)  were harvested either into ice cold Ca2+Mg2+-free PBS (CMF-PBS) containing 0.2mM EDTA for tissue separation, or into ice-cold Tyrode's saline for tissue dissection. To separate the endocardium from ventricular tissue, 10µg/ml hyaluronidase from bovine testes was used. HH12 hearts were separated from surrounding splanchnic mesoderm, foregut and anterior-intestinal portal (AIP) endoderm. Dissected tissues were collected and pooled in 200ul of ice-cold RNA stabilisation solution, RNAlater and stored at 4°C. RNA was isolated using RNAqueous Micro-kit according to the manufacturer's instructions. DNA was removed by applying DNase treatment. The RNA quality was measured by using Agilent 2200 TapeStation Libraries were constructed from 100ng total RNA using the NEBNext® Ultra™ II Directional RNA Library Prep Kit with NEBNext® Poly(A) mRNA Magnetic Isolation Module according to the manufacturer's instructions. Libraries to be multiplexed in the same run were pooled in equimolar quantities
attributes.tag: Experimental Factor: organism part
attributes.value: germ wall
attributes.tag: Experimental Factor: sampling site
attributes.value: not applicable