Formalin-fixed, paraffin-embedded (FFPE) gallbladder tissue specimens were obtained from 98 patients in total (n = 31 GS; n = 35 Dys; n = 32 GBC). RNA was extracted from FFPE sections using the AllPrep FFPE kit following Qiagen’s recommendations, and RNA quality was controlled (High Sensitivity Genomic DNA, Advanced Analytical, United States, and FFPE quality control kits, Illumina).
The NEBNext Small RNA kit (NEB) was used to produce RNA sequencing libraries, which were sequenced on the HiSeq 2500 platform (Illumina, San Diego, CA, USA) to an average depth of 18 M reads per sample. The applied RNA sequencing protocol has been previously described in detail [18 (link)]. Briefly, our protocol enabled us to capture lncRNA mapped fragments in the size range up to 47 base pairs. First, reads from the HiSeq 2500 platform were adapter-trimmed (AdapterRemoval v2.1.7) [19 (link)]. Then, adapter-trimmed reads were mapped to the human genome (hg38) by a Bowtie2 v2.2.9 aligner [20 (link)]. HTSeq was used to count reads mapped to lncRNA regions in GENCODE v26 annotations [21 (link),22 (link)].
The NEBNext Small RNA kit (NEB) was used to produce RNA sequencing libraries, which were sequenced on the HiSeq 2500 platform (Illumina, San Diego, CA, USA) to an average depth of 18 M reads per sample. The applied RNA sequencing protocol has been previously described in detail [18 (link)]. Briefly, our protocol enabled us to capture lncRNA mapped fragments in the size range up to 47 base pairs. First, reads from the HiSeq 2500 platform were adapter-trimmed (AdapterRemoval v2.1.7) [19 (link)]. Then, adapter-trimmed reads were mapped to the human genome (hg38) by a Bowtie2 v2.2.9 aligner [20 (link)]. HTSeq was used to count reads mapped to lncRNA regions in GENCODE v26 annotations [21 (link),22 (link)].
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