SRA

The hypothalamic medio-basal region was recovered by dissection from one male of the E- lineage of Coturnix japonica (doi: 10.1037/0735-7036.105.1.15) that was bred under 16L/8D light condition. The dissected region was frost in liquid nitrogen then kept at -80°C until extraction. Total RNAs were purified using Trizol. PacBio Iso-Seq library preparation and sequencing were performed by the INRAE platform Gentyane (http://gentyane.clermont.inrae.fr). cDNAs were prepared using the kit NEBNext Single Cell/Low Input cDNA Synthesis and Amplification Module, New England Biolabs(ref.E6421S/L) with some buffer modifications done in order to limit the impact of G-quadruplexes that are Na+ and K+ dependent on their assembly. For the reverse transcriptase (RT) and the Q5 amplification steps, CsCl was used to replace KCl in the 5X Single Cell RT Buffer (250 mM Tris-HCI (pH 8.3), 250 mM CsCl2,15 mM MgCl2, 50 mM dithiothreitol) and 5X Q5 Buffer (250 mM Tris-HCl (pH 8.3 @ 25°C), 250 mM CsCl2, 20 mM MgCl2, 100 mM (NH4)2SO4, 10 mM DTT). The library m64071 was made with just a buffer modification for the RT step while the library m64243 had both steps done with buffer modification. Because RT is less efficient with CsCl2, each library was made from 2 pooled RT reactions made using for each 500 ng total RNA, and a modified procedure consisted of a denaturation step of 10 min at 70°C, a RT extension step of 105 min at 42°C, and a final extension step of 30 min at 42°C, after adding Template switching oligo (TSO). The amplification with the Q5 was done with 15 cycles and an extension time of 5 min per cycle. Sequencing was performed on the Sequel II system at the Gentyane Genomic Platform (INRAE Clermont-Ferrand, France) with Sequel II Sequencing kit 3.0.