TY - JOUR
T1 - RNA sequencing and swarm intelligence–enhanced classification algorithm development for blood-based disease diagnostics using spliced blood platelet RNA
AU - Best, Myron G.
AU - in ’t Veld, Sjors G. J. G.
AU - Sol, Nik
AU - Wurdinger, Thomas
AU - In 't Veld, Sjors G J G
PY - 2019/4/1
Y1 - 2019/4/1
N2 - Blood-based diagnostics tests, using individual or panels of biomarkers, may revolutionize disease diagnostics and enable minimally invasive therapy monitoring. However, selection of the most relevant biomarkers from liquid biosources remains an immense challenge. We recently presented the thromboSeq pipeline, which enables RNA sequencing and cancer classification via self-learning and swarm intelligence–enhanced bioinformatics algorithms using blood platelet RNA. Here, we provide the wet-lab protocol for the generation of platelet RNA-sequencing libraries and the dry-lab protocol for the development of swarm intelligence–enhanced machine-learning-based classification algorithms. The wet-lab protocol includes platelet RNA isolation, mRNA amplification, and preparation for next-generation sequencing. The dry-lab protocol describes the automated FASTQ file pre-processing to quantified gene counts, quality controls, data normalization and correction, and swarm intelligence–enhanced support vector machine (SVM) algorithm development. This protocol enables platelet RNA profiling from 500 pg of platelet RNA and allows automated and optimized biomarker panel selection. The wet-lab protocol can be performed in 5 d before sequencing, and the algorithm development can be completed in 2 d, depending on computational resources. The protocol requires basic molecular biology skills and a basic understanding of Linux and R. In all, with this protocol, we aim to enable the scientific community to test platelet RNA for diagnostic algorithm development.
AB - Blood-based diagnostics tests, using individual or panels of biomarkers, may revolutionize disease diagnostics and enable minimally invasive therapy monitoring. However, selection of the most relevant biomarkers from liquid biosources remains an immense challenge. We recently presented the thromboSeq pipeline, which enables RNA sequencing and cancer classification via self-learning and swarm intelligence–enhanced bioinformatics algorithms using blood platelet RNA. Here, we provide the wet-lab protocol for the generation of platelet RNA-sequencing libraries and the dry-lab protocol for the development of swarm intelligence–enhanced machine-learning-based classification algorithms. The wet-lab protocol includes platelet RNA isolation, mRNA amplification, and preparation for next-generation sequencing. The dry-lab protocol describes the automated FASTQ file pre-processing to quantified gene counts, quality controls, data normalization and correction, and swarm intelligence–enhanced support vector machine (SVM) algorithm development. This protocol enables platelet RNA profiling from 500 pg of platelet RNA and allows automated and optimized biomarker panel selection. The wet-lab protocol can be performed in 5 d before sequencing, and the algorithm development can be completed in 2 d, depending on computational resources. The protocol requires basic molecular biology skills and a basic understanding of Linux and R. In all, with this protocol, we aim to enable the scientific community to test platelet RNA for diagnostic algorithm development.
KW - Biomarkers/blood
KW - Blood Platelets/chemistry
KW - Computational Biology/methods
KW - DNA, Complementary/analysis
KW - High-Throughput Nucleotide Sequencing/methods
KW - Humans
KW - RNA Splicing
KW - RNA, Messenger/analysis
KW - Sequence Analysis, RNA/methods
KW - Support Vector Machine/statistics & numerical data
UR - https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=85063297725&origin=inward
UR - https://www.ncbi.nlm.nih.gov/pubmed/30894694
U2 - https://doi.org/10.1038/s41596-019-0139-5
DO - https://doi.org/10.1038/s41596-019-0139-5
M3 - Article
C2 - 30894694
SN - 1754-2189
VL - 14
SP - 1206
EP - 1234
JO - Nature Protocols
JF - Nature Protocols
IS - 4
ER -