Name: GSM7937318
Instrument: Illumina NovaSeq 6000
Strategy: RNA-Seq
Source: TRANSCRIPTOMIC
Selection: cDNA
Layout: PAIRED
Construction protocol: Embryo total RNA was extracted from each biological replicate using the PicoPure RNA isolation kit (Thermo Fisher Scientific) with DNAse treatment (Qiagen) as per the manufacturer's protocol. RNA was eluted in 15 µL of nuclease-free water and kept at -80°C for subsequent library preparation. RNA concentration and integrity were evaluated using an Agilent 4200 TapeStation (Agilent Technologies) with the High Sensitivity RNA ScreenTape Assay (Agilent Technologies). RNA Integrity Number (RIN) ranged from 5.0 to 8.3 RIN with an average of 6.2 ± 1.4 RIN and RNA concentration ranged from 308 to 556 pg/µL with an average of 414 ± 113.6 pg/µL. Library preparation was performed for each biological replicate. Enrichment of mRNA was conducted using the NEBNext Poly(A) mRNA Magnetic Isolation Module (E7490, New England Biolabs, USA) and libraries were prepared using the NEBNext Ultra II RNA Library kit for Illumina (E7770, E7775, New England Biolabs, USA). The library preparation protocol was followed as recommended by the manufacturer's guidelines for 10 ng – 1μg of DNA-free total RNA, with 15 cycles for the PCR enrichment of adaptor-ligated DNA and Illumina index primers (Illumina). Purified libraries were eluted in 23 μL of (0.1X) TE buffer and stored at -20°C for future analysis. The quality of the 6 libraries were verified using Agilent bioanalyzer (4200 TapeStation, Agilent Technologies) with the High Sensitivity D1000 ScreenTape Assay (Agilent Technologies) before libraries were pooled in equimolar proportion and sequenced with NovaSeq 6000 using paired-end 100bp protocol.