SRA

Ustilago maydis cells over-expressing 3xFLAG-UmaTERT (telomerase reverse transcriptase) lysed and immunoprecipitation (IP) performed with anti-FLAG antibodies to IP telomerase. Co-immunoprecipitated RNA extracted and subject to illumina next-generation sequencing (NGS) followed by computational screening of TR candidates from NGS data and experimental validation of top candidate. Nanopore long read sequencing libraries were generated using total RNA from wild type cells to characterize TR precursor transcripts or from recombinant Ustilago maydis cells expressing a mutated TR precursor to determine maturation mechanism of TR. For illumina library, Illumina Scriptseq v2 RNA-seq library preparation kit was used and sequenced on a Nextseq 500. For Nanopore long read libraries, PCR-cDNA sequencing kit (SQK-PCS109 - Oxford Nanopore Technologies) was used with additional nested PCR steps to enrich targets and sequenced on a minION device. All reads subject to adapter trimming and quality control using tools specific for each technology.