| description |
SCD6 and Rasputin are two RNA binding proteins of the parasite Leishmania braziliensis. The main interest is to know at which RNA they bind. To achieve this task, SCD6-RNA and Rasputin-SCD6 complexes were immunoprecipitated,using antibodies against each of these proteins, starting from a parasite extract (cells lysed by sonication). In parallel, immunoprecipitation with the pre-immune sera was carried out as control of non specific binding (there are two control, preRasputin and preSCD6). Once the complexes were obtained, proteins were removed and the RNA was concentrated by ethanol precipitation. As the amount of RNA was undetectable,the TransPlex® Whole Transcriptome Amplification Kit (Sigma-Aldrich) system1 were used to generate cDNA and then amplified by PCR to obtain enough mass of cDNA to perform further steps of NGS analysis. There is no accurate data on the nucleotide sequence that the kit incorporates in PCR stage. |