GSM8563957: TCR1; Hakea laurina; RNA-Seq
The total RNA from frozen roots was extracted using the RNeasy Plant Mini Kit (Qiagen, Hilden, Germany) with modifications for woody plant tissues. Briefly, 50 mg of frozen sample was homogenized with 1 mL of extraction buffer containing 4 M guanidine isothiocyanate, 0.2 M sodium acetate at pH 5.0, 25 mM EDTA, 2.5% (w/v) PVP-25 and 1% β-mercaptoethanol. The homogenates, to which 100 µL of 20% (w/v) sarkosyl was added, were incubated at 70˚C for 10 min with occasional vortexing. All lysates were transferred onto a QIAshredder Spin Column, followed by the standard protocol. The cDNA libraries for RNA-Seq were prepared using an NEBNext Ultra ll Directional RNA Library Prep Kit for Illumina, NEBNext Multiplex Oligos for Illumina, and NEBNext Poly(A) mRNA  Magnetic Isolation Module (New England Biolabs Japan, Inc., Tokyo, Japan) following manufacturer's protocols.