MicroRNAs for the identification of exposure to particulate matter 2.5 (pm2.5) and methods of identification using the same
A kind of use and aspect technology, applied in the field of microRNA, can solve the problems of uncertainty, PM2.5 research limitation composition, PM2.5 risk assessment data shortage, etc.
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Embodiment 1
[0116] Example 1: Collection and extraction of PM2.5
[0117] Selection of collection locations
[0118] To screen biomarkers for the identification of PM2.5 exposure, the roof of the Korea Institute of Science and Technology (KIST) gymnasium (Hawolgok-dong, Sungbuk-gu, Seoul) was chosen as the collection site for PM2.5 collection. The stadium is adjacent to Naebu Expressway near Wolgok Station in Sungbuk-gu, where the traffic volume is as high as 70,000 vehicles on weekdays in November and 69,000 vehicles on weekdays in December (2011) . For in vitro testing, considerable amounts of PM2.5 are required. Since the low-volume air sampler can only obtain a few mg of PM2.5 after running for 24 hours, the low-volume air sampler is not suitable for the collection of PM2.5. Therefore, for in vitro testing of PM2.5, high volume air samplers are used to facilitate the collection of high volume particles. PM2.5 specific inlet (Tisch Instrument Company) collected PM2.5.
[0119] ...
Embodiment 2
[0125] Example 2: Selection of PM2.5 exposure model and acquisition of mouse lung tissue samples
[0126] PM2.5 Exposure Model
[0127] PM2.5 is suspended particulate matter in the air and is one of the harmful substances in the environment. Once particles enter the lungs, typical particle-induced inflammation will be induced. Based on previous reports, the BALB / C model was used to measure such responses with high sensitivity. BALB / C are susceptible to chronic pneumonia and have high sensitivity to radiation. This animal is relatively tame and docile, and shows a lower incidence of breast cancer. Therefore, this animal was finally selected as a suitable model.
[0128] Specifically, the PM2.5 collected in was divided into two different groups, namely water-soluble group and organic-soluble group. The mouse model was also divided into two groups, one group was exposed to PM2.5 and the other group was not exposed to PM2.5. The exposed group was exposed to either low doses...
Embodiment 3
[0151] Example 3: Microarray Assays
[0152] Isolation of target microRNA and labeling with fluorescent substances
[0153] RNA was extracted from blood samples of PM2.5-exposed and PM2.5-unexposed groups using trizol. The extracted RNA was purified using RNeasy Mini kit (Cat NO.74106, Qiagen). During RNA purification, genomic DNA was removed using RNase-free DNase Set (Qiagen, USA). The concentration of each total RNA was measured with a NanoDrop ND 1000 spectrophotometer (NanoDrop Technologies Inc., USA). The quality of RNA was measured using an Agilent 2100 Bioanalyzer.
[0154] Preparation of labeled microRNA
[0155] For the oligonucleotide microarray assay, the total RNA obtained from the PM2.5-exposed group and the PM2.5-unexposed group of Example was labeled with a fluorescent substance. Labeling was performed using the miRNA complete labeling and hybridization kit (Agilent) following the manufacturer's protocol. The obtained total RNA (100 ng) was mixed with ...
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