title: NextSeq 500 sequencing; Transcriptome analysis of human brain microvascular endothelial cells response to Borrelia burgdorferi and its antigen Erp23 using RNA-seq
design: Transcriptome analysis of human brain microvascular endothelial cells response to Borrelia burgdorferi and its antigen Erp23 using RNA-seq
library_construction_protocol: Human brain microvascular endothelial cells (hBMECs) were cultured in 6-well plates then incubated either with live Borrelia burgdorferi or recombinant Erp23  or culture medium only (non-induced control) for 6 hours.  Human BMECs (hBMEC/D3 cell line), were acquired from Merck/Millipore (Prague, Czech Republic). Cells were cultured in 6-well cell culture plates covered with collagen type I (Sigma, USA) and endothelial cell growth basal medium (EBM-2; Lonza,UK; Appendix 1). Cells were incubated at 37°C in a humid atmosphere in the presence of 5% CO2.  B. burgdorferi (strain Pib) were cultured in BSK-2 medium. Bacteria were grown at 37°C.   Human brain endothelial cells cultured in 6-well plates were incubated either with live Borrelia burgdorferi (MOI 0.5,bacterial cells were washed with minimal essential medium, Sigma) or recombinant Erp23(27 μg/well). Nontreated cells were used as negative control,  for 6 hours at  37°C in the presence of 5% CO2. mRNA from human brain microvascular endothelial cells was isolated using RNeasy Mini Kit (Qiagen, Germany) according to manufacturer's instructions. 250 ng of RNA were reverse transcribed with oligodT primers using QuantSeq 3′ mRNA-Seq Library Prep Kit (Lexogen, Austria).
attributes.tag: Experimental Factor: infect
attributes.value: Borrelia burgdorferi
attributes.tag: Experimental Factor: stimulus
attributes.value: none
attributes.tag: Experimental Factor: time
attributes.value: 6