GSM9108393: mSI_org_Lkb1HomBud_APCKO_1; Mus musculus; RNA-Seq
Organoids were lysed in RLT lysis buffer (Qiagen) 3-5 days after passaging. Biological triplicates were employed. To isolate RNA from samples, the “miRNA CT 400” protocol was run on a QIAsymphony isolation robot using the QIAsymphony RNA Kit (931636) on each sample in 400ul of the RLT Plus lysis buffer (1053393). RNA quality was checked with the Agilent Fragment Analyzer 5300 system using the RNA Kit (15nt) (Cat. DNF-471-1000). And RNA quantity was measured with the Invitrogen™ Qubit ™ Fluorometer using the Qubit RNA HS Assay Kit (Cat. Q32855). 100ng of total RNA was used to prepare TruSeq Stranded mRNA libraries (Cat. 20020594) following the manufacturers protocol, with full (WT, Lkb1+/- and Lkb1-/- organoids) or half (Lkb1mut + Apc-/- and Lkb1mut + KrasG12D organoids) the reagents volume. After the library preparation libraries were checked with the Fragment Analyzer system dsDNA 910 Reagent Kit (35-1500bp) (Cat. DNF-910-K1000) and with Qubit dsDNA HS Assay Kit (Cat. Q32854). Sample libraries were pooled equimolar.