organism: Yellow fever virus
description: Protocols: Yellow Fever infectious clones (IC) were generated by site-directed mutagenesis (SDM) of either pANCR 17D (IC) or pANCR Asibi IC as a template. QuickChange XLII SDM kit (Strategene) along with mutagenic primer sets were used to generate infectious clones as per the manufacturer's protocol. Stock cDNA YF IC were amplified in E. coli (MC1061) and plasmid harvested by large-scale extraction. Full-length YF RNA was generated by in vitro transcription of purified cDNA using SP6-Scribe standard RNA in vitro transctiption kit (Cellscript) according to the manufacturer's protocol; the reaction was incubated at 37oC for 2.5 hours. The entire reaction was electroporated into 6.7x106 monkey kidney Vero cells. Cell supernatant was collected when infected cells showed 80% cytopathic effect (CPE). RNA from the YF ICs were extracted using QIAamp Viral RNA mini kit (Qiagen), following the manufacturer's guidelines. NGS libraries were constructed using TruSeq RNA v2 kit as recommended by the manufacturer.
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2018-10-16T17:02:47Z
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2018-03-20T16:28:12Z
attributes.tag: External Id
attributes.value: SAMEA104708356
attributes.tag: INSDC center name
attributes.value: Department of Microbiology & Immunology, University of Texas Medical Branch
attributes.tag: INSDC first public
attributes.value: 2018-10-16T17:02:47Z
attributes.tag: INSDC last update
attributes.value: 2018-03-20T16:28:12Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-6614:Sample 3
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: genotype
attributes.value: NS4B-M95I substitution
attributes.tag: growth condition
attributes.value: grown in monkey kidney Vero cells
attributes.tag: sample name
attributes.value: E-MTAB-6614:Sample 3
attributes.tag: scientific_name
attributes.value: Yellow fever virus
attributes.tag: strain
attributes.value: 17D-204