MicroNA chip experiment

microRNA chip experiment flow 1. Sample RNA extraction
a. Experiment objects for tissue samples, take appropriate amount (50-100 mg) fresh tissue sample or orthimate tissue sample, use BiopulverizertM frozen crush Tissue, add 1 ml of RNA extract reagent Trizol (Invitrogen), and extracted RNA after using mini-beater-16 homogenous.
b. Experimental objects are cell samples, each sample takes 1 u0026 Times; 106 ~ 1 u0026 Times; 107 cells, add 1 ml of RNA extract reagent Trizol (sample is a wall cell Each 10 cm2 petri dish Trizol is 1 mL), and the RNA is extracted after cleavage. 2. RNA quality detection
a. The RNA was measured using a NanoDrop to measure the absorption value of 260 nm, 280 nm, 230 nm, to calculate the concentration and evaluate the purity.
b. Determinate agarose gel electrophoresis with formaldehyde electrophoresis reagents, detect RNA purity and integrity.
c. Provide RNA QC report.
Note: RNA samples for chip detection are high quality, complete, no RNASE pollution (degraded samples cannot be used for markers and chip detection), no Genomic pollution. 3. Preparation of fluorescent label probes
uses mircury u0026 trade; array power tag kits, HY3 u0026 TRADE; or HY5 u0026 TRADE; fluoride group miRNA It can be obtained for fluorescent probes for hybridization with the chip. 4. Chip hybridization
Use the MAUI hybridizer to hybridize to Mircury u0026 Trade; chip under standard conditions. 5. Image acquisition and data analysis
u0026 nbsP; use the GenePix 4000B chip scanner to scan the fluorescence intensity of the chip, and convert the experimental results into digital data storage, use the supporting software to analyze the original data.
6. Provide experiment report u0026 mdash; u0026 mdash; includes detailed experimental methods and chip experiment data and chart a. Chip scan

123] b. Operating instructions (containing RNA quality inspection report)
c. Chip data
u0026 nbsp ; Data summary table (Excel format, complicated probe location, probe name, raw signal value, correction signal value, standardized signal value, sample room microRNA expression quantity)

Differentially express MicroRNA's data sheet (Excel format, microRNA, which contains 2 times, up-regulated microRNA and Double-regulation of microRNA)
d. further data analysis [ 123] Scatter Plot (mainly for monochrome repeat experiment data)
layered cluster (When performing multiple sample experiments, clustering is carried out according to the degree of gene expression, thereby categoring multiple samples)
Then calculate the CV value, p value, and do the Volcano PLO

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