Severe acute respiratory syndrome-related coronavirus
Protocols: Lysis of infected cell monolayer in trizol. Cells were incubated with 200 uL Trizol at room temperature for 15 minutes before collection into 2 mL tubes. Trizol extraction using zymo directzol kit per manufacturer's protocol. a region surrounding the N.iORF3 TRS-B sequence was amplified using Platinum SuperFi II DNA Polymerase (Invitrogen) with primers N3geno_FWD and N3geno_REV which contained 5′ Illumina adapter sequences. PCR cycling conditions were 95 ºC for 5 minutes, followed by 20 cycles of 95 ºC for 30 s, 60 ºC for 30 s, and 72 ºC for 20 s, with a final extension of 72 ºC for 5 minutes. Products were cleaned up using Ampure XP beads (Beckman Coulter) at a ratio of 1.8 x. 1 µL of each sample was taken into a PCR reaction with 5 µL of NEB Q5 High-Fidelity DNA Polymerase 2X Master Mix, 1 µL of NXT-IDT Primer Mix 10 µM unique dual index and 3 µL of H2O. PCR cycling conditions were 95 ºC for 3 minutes, followed by 10 cycles of 95 ºC for 30 s, 55 ºC for 15 s, and 72 ºC for 30 s, with a final extension of 72 ºC for 5 minutes. Libraries were cleaned up using Ampure XP beads at a ratio of 0.8 x. Libraries were quantified using Promega Quantifluor reagents, plate reader and Agilent Tapestation.
ENA first public
2024-01-01
INSDC center name
The Francis Crick Institute
INSDC status
public
Submitter Id
E-MTAB-14680:Alpha-WTvssilTRS_8hpi_1
broker name
ArrayExpress
collection date
not collected
geographic location (country and/or sea)
not collected
isolate
not applicable
sample name
E-MTAB-14680:Alpha-WTvssilTRS_8hpi_1
scientific_name
Severe acute respiratory syndrome-related coronavirus