Delivery Standards for Customized Detection Services of Circular RNA Chips

Customized detection service for circular RNA chips

Introduction to Experimental Techniques: Circular RNA(ircRNA) is a new type of RNA that differs from traditional linear RNA. It has a closed circular structure and is widely present in the eukaryotic transcriptome. Most circular RNAs are composed of exon sequences, which are conserved in different species and exhibit tissue-specific and developmental stage specific expression. Due toCircular RNABeing insensitive to nucleases, circular RNA is more stable than linear RNA, which gives it a significant advantage in the development and application of novel diagnostic markers. Recent studies have shown that circular RNA acts as a miRNA sponge in different species, known as competitive endogenous RNA (ceRNA), which can competitively bind to miRNA. The interaction between miRNAs associated with diseases indicates that circular RNAs play a crucial role in regulating diseases.

Experimental operation process:

1. Sample RNA extraction

2. RNA quality testing

① Measure the absorption values of RNA at 260nm, 280nm, and 230nm using Nanodrop spectrophotometer to calculate concentration and evaluate purity.

② Formaldehyde electrophoresis reagent was used for denatured agarose gel electrophoresis to detect the purity and integrity of RNA.

③ Provide RNA QC report.

3. RNase R processing

4. Synthesis and labeling of cDNA/mRNA samples

5. The labeling efficiency quality testing uses Nanodrop to detect the fluorescence labeling efficiency, and the labeling efficiency is qualified to ensure the reliability of subsequent chip experimental results.

6. Chip hybridization

Hybridize the labeled probes with high-density genomic chips under standard conditions.

7. Image acquisition and data analysis

Use GenePix 4000B chip lunar scanner to scan the fluorescence intensity of the chip, and convert the experimental results into digital data for storage. Use supporting software to analyze and calculate the raw data.

8. Provide experimental report

① Scanning Image: Cy3 Fluorescence Scanning Image;

② Scatter Plot: A scatter plot where the X-axis represents the Control signal values and the Y-axis represents the Experiment signal values, indicating the overall distribution trend of the signal data;

③ Raw Data: The raw data of the scanning fluorescence signal intensity of the probe;

④ Normalized Data: numerical values standardized by statistical methods;

⑤ P-value: T-test statistical analysis of significantly differentially expressed genes. The smaller the p-value, the more significant the differential expression of this circRNA between the two groups of samples

⑥ Significant Up&Down Regular CircRNA List: The list of significantly differentially expressed circRNAs includes Fold Change, p-valuel, and detailed annotation information for circRNAs (corresponding linear mRNA and miRNA binding sites).

The customer provides:

1. The experimental subjects are tissue samples. Take an appropriate amount (50-100mg) of fresh tissue samples or correctly stored tissue samples, and use them

BioPulverizer TM frozen crushed tissue was added with 1m RNA extraction reagent TRIzol (Invitrogen) and homogenized with Mini-Bad-Bater-16 to extract RNA.

2. The experimental subjects are cell samples, with 1x106~1x107 cells taken from each sample, and 1m of RNA extraction reagent TRIzol added (the sample is adherent cells, and the amount of TRIzol used per 10cm2 culture dish is 1ml). RNA is extracted after lysis.

Delivery standards:

1. Chip detection data

2. Complete experimental operating procedures, experimental reports (including software analysis results), and experimental materials

Experimental outsourcing service: