Transcriptomic analysis of Morchella sextelata
First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase(RNaseH). Second strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNaseH. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3 ends of DNA fragments, NEBNext adaptor with hairpin loop structure were ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 250-300 bp in length, the library fragments were purified with AMPure XP system (Beckman Coulter, Beverly, USA).Then 3 l USER Enzyme (NEB, USA) was used with size-selected, adaptor-ligated cDNA at 37C for 15 min followed by 5 min at 95 C before PCR. Then PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers and Index (X) Primer. At last, PCR products were purified (AMPure XP system) and library quality was assessed on the Agilent Bioanalyzer 2100 system.