Using QIAzol lysis reagent (QIAGEN), messenger RNA (mRNA) was extracted from the middle ear mucosa of the rats in both the control and exposure groups. Complementary DNA (cDNA) was synthesized using Moloney murine leukemia virus (Promega). To determine the expression of IL-1ß, IL-6, TNF-α, and vascular endothelial growth factor (VEGF), RT-PCR was performed twice using the 2X real-time PCR Master mix SYBR Green (Thermo Fisher Scientific), according to the manufacturer’s instructions. Glyceraldehyde 3-phosphate dehydrogenase values were used to normalize gene expression. The PCR primers have been referenced in a similar study (Table 1 ) [18 (link)].
Protocol detail
Quantifying Inflammatory Markers in Rat Middle Ear
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Cdna
Gene expression
Glyceraldehyde 3 phosphate dehydrogenase
Messenger rna
Middle ear
Moloney murine leukemia virus
Mucosa
Primers
Promega
Rats
Real time pcr
Rt pcr
Sybr green
Tnf α
Vascular endothelial growth factor
Corresponding Organization :
Other organizations : Pusan National University Yangsan Hospital, Pukyong National University, Inje University Haeundae Paik Hospital, Pusan National University Hospital
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Variable analysis
independent variables
- Exposure group
- Expression of IL-1ß
- Expression of IL-6
- Expression of TNF-α
- Expression of vascular endothelial growth factor (VEGF)
- Control group
- Positive control: Not specified
- Negative control: Not specified
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