Sample 19
Equus caballus
Protocols: Synovial fluid samples were obtained from both middle carpal joints prior to (day 0), day 28 and 70 days after surgery. Horses were sedated with detomidine (0.01 mg/kg, Domosedan Vet, Orion Pharma Animal Health, Copenhagen, Denmark) and butorphanol tartrate (0.01 mg/kg, Dolorex, MSD Animal Health, Copenhagen, Denmark), and the dorsal surface of the carpi prepared aseptically by chlorhexidine scrubbing and alcohol swabbing. Synovial fluid was aspirated using a 19 gauge 40 mm needle and immediately transferred into ethylene diamine tetra acetic acid (EDTA) tubes (BD Vacutainer, BD A/S, Albertslund, Denmark). Tubes were inverted carefully 5-10 times and samples were further processed as detailed in Table 1 within one hour and stored at -80 ‏⁰C until small RNA sequencing and RT-qPCR analysis of miRNA expression (performed within 6 months). 500 µl SF were treated with a 1ug/µl hyaluronidase (HA) solution to reduce viscosity as preciously described [12]. RNA was extracted from SF using miRNeasy serum/plasma Advanced Kit (Qiagen, Vedbæk, Denmark) according to the manufacturer's instructions. Samples underwent on-column DNase digestion with RNase free DNase sets (Qiagen, Vedbæk, Denmark) and RNA yield was determined by spectrophotometry (Nano Drop ND-1000 spectrophotometer, Saveen and Werner AB, Limhamn, Sweeden). This project utilized the NEBNext® Small RNA Library Prep Set for Illumina® (Multiplex Compatible). The protocol was adapted to accommodate the low input of sample by dilution of adapters 1 in 5 and extra PCR cycles.
sampling site
midcarpal joint
isolate
not applicable
ENA-FIRST-PUBLIC
2023-03-01T00:16:44Z
individual
H8
organism
Equus caballus
organism
Equus caballus
ENA-LAST-UPDATE
2023-03-01T00:16:44Z
sex
male
scientific_name
Equus caballus
common name
horse
organism part
synovial fluid
age
5
developmental stage
mature
ENA-FIRST-PUBLIC
2023-03-01
ENA-LAST-UPDATE
2023-03-01