DNA and RNA were extracted from the parasite pellets with the AllPrep DNA/RNA Mini Kit (Qiagen). The RNA integrity was then verified with the RNA 6000 Nano Kit using a Bioanalyzer 2100 (Agilent). DNA was quantified with the Qubit dsDNA BR assay (Thermo Fisher Scientific) and RNA with the Qubit RNA HS Assay.
Library preparation and sequencing of the DNA samples were performed at the Beijing Genomics Institute (BGI). Libraries were prepared with the TruSeq Nano DNA HT sample prep kit (Illumina) and 2 x 151 bp sequenced on the Illumina HiSeq 4000 platform. RNA sequencing libraries were prepared using the Spliced leader sequencing (SL-Seq) method as described in Cuypers et al. (2017) [16 (link)]. This protocol makes use of the presence of the affixed 39 nucleotide sequence spliced-leader (SL) that is present at the 5’ end of all functional Leishmania mRNAs. RNAs containing a SL are selectively amplified with the protocol, and adapters required for Illumina sequencing are ligated. The SL-Seq libraries were 1 X 50 bp sequenced with the HiSeq 1500 platform of the Center of Medical Genetics Antwerp (Belgium).
Library preparation and sequencing of the DNA samples were performed at the Beijing Genomics Institute (BGI). Libraries were prepared with the TruSeq Nano DNA HT sample prep kit (Illumina) and 2 x 151 bp sequenced on the Illumina HiSeq 4000 platform. RNA sequencing libraries were prepared using the Spliced leader sequencing (SL-Seq) method as described in Cuypers et al. (2017) [16 (link)]. This protocol makes use of the presence of the affixed 39 nucleotide sequence spliced-leader (SL) that is present at the 5’ end of all functional Leishmania mRNAs. RNAs containing a SL are selectively amplified with the protocol, and adapters required for Illumina sequencing are ligated. The SL-Seq libraries were 1 X 50 bp sequenced with the HiSeq 1500 platform of the Center of Medical Genetics Antwerp (Belgium).
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