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| Product Name | Silica-Membrane Total RNA Purification Kit |
| Catalog No. | NATR-HMM-0112 |
| Description | A spin column-based kit for the rapid purification of total RNA from cultured cells, animal tissues, and biological fluids. The guanidine-based lysis and ethanol precipitation combined with on-column DNase digestion provides high-quality RNA free of genomic DNA contamination. |
| Intended Use | Purification of total RNA from a variety of biological samples for gene expression analysis by RT-qPCR, microarray hybridization, Northern blotting, and RNA-seq library construction. |
| Principle / Technology | Cells or tissues are disrupted and homogenized in a guanidine isothiocyanate-based lysis buffer containing β-mercaptoethanol to inactivate RNases. Ethanol is added to adjust binding conditions, and the lysate is passed through a silica membrane where RNA selectively binds. On-column DNase I treatment removes co-purified genomic DNA, and after wash steps, pure total RNA is eluted in RNase-free water. |
| Detection Method | UV spectrophotometry; agarose gel electrophoresis with formaldehyde denaturation; Bioanalyzer electrophoretic analysis for RNA Integrity Number (RIN); RT-qPCR amplification of reference genes |
| Sample Type | Cultured mammalian cells (adherent and suspension), fresh and frozen animal tissues, white blood cells, yeast, bacteria (with appropriate pretreatment), plant tissues |
| Performance Range / Specifications | RNA binding capacity per column: 100 μg; typical yield from 10^7 HeLa cells: 40–80 μg; from 30 mg mouse liver: 60–120 μg; A260/A280 ratio: 1.9–2.1 |
| Sensitivity / LOD | Recovers total RNA from as few as 100 cells with micro-isolation protocol; detects low-abundance transcripts (<10 copies per cell) by RT-qPCR |
| Specificity | Recovers RNA species >200 nucleotides; small RNAs (<200 nt) recovered with modified protocol; no residual genomic DNA detectable by PCR following on-column DNase treatment |
| Reaction Conditions / Protocol | Homogenize sample in lysis buffer (350–600 μL), add equal volume 70% ethanol, mix, transfer to spin column, centrifuge 15 seconds, apply DNase I enzyme mixture to membrane, incubate 15 minutes at room temperature, wash with wash buffer RW1, wash twice with buffer RPE, dry spin, elute with 30–100 μL RNase-free water; total time approximately 30 minutes (excluding DNase incubation) |
| Components / Formulation | Buffer RLT (guanidine isothiocyanate, Tris-HCl, EDTA), Buffer RW1 (guanidine hydrochloride, Tris-HCl, ethanol), Buffer RPE concentrate (Tris-HCl, NaCl, ethanol), RNase-free water, DNase I (lyophilized), RDD buffer for DNase reconstitution, spin columns with silica membrane and collection tubes |
| Storage Conditions | Lyophilized DNase I and buffers stored at 2–8°C after reconstitution; all other components at room temperature; protect Buffer RLT from light |
| Shelf Life | 24 months for columns and buffers; 12 months for reconstituted DNase I at 2–8°C |
| Package Specifications | 50 and 250 preparation kits; all columns, tubes, and reagents included |
| Product Form | Liquid buffers; silica membrane spin columns; lyophilized DNase I with reconstitution buffer |
| Quality Control | Each lot validated with HeLa cell RNA: yield ≥40 μg per 10^6 cells; A260/A280: 1.9–2.1; gDNA-free verified by human genomic PCR with GAPDH intron-spanning primers; RIN ≥8.0; functional testing by SYBR Green RT-qPCR efficiency 90–110% for six reference genes |
| Key Features | On-column DNase digestion eliminates separate DNase treatment step; RNase-free buffers and plasticware eliminate degradation risk; rapid protocol under 30 minutes; high RNA binding capacity for concentrated samples; validated with QIAGEN, Agilent, and Roche downstream platforms |
For research use only, not for clinical use.
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