PacBio IsoSeq sequencing for Glycine soja W05
Multiple tissues of Glycine soja W05 were collected for PacBio IsoSeq sequencing. Embryo was collected after 24 hours of germination. Cotyledons and hypocotyl were collected 3 days after sowing, while root, apical buds and stems were collected after 10 days after sowing. Flower, 7-day pods, 14-day pods, 14-day seeds, 40-day pods, 40-day seeds were collected at reproductive stages. Total RNA was extracted from these tissues using RNAiso Plus reagent. In addition, nodules and remaining root were collected from seedlings 28 days post inoculation with Sinorhizobium fredii strains CCBAU 45436. RNA sample was extracted from these two samples using TRIzol reagent. All the RNA samples were mixed together and four SMRTbell libraries were constructed with size selection of 1-2Kb, 2-3Kb, 3-6Kb and 5-10Kb using the BluePippin. SMRTbell libraries were sequenced on PacBio RSII platform with P6-C4 chemistry.