Quick osseous tissue RNA extraction kit

A bone tissue and kit technology, applied in recombinant DNA technology, DNA preparation, etc., can solve the problems of low bone cell density, inability to conduct experiments, and RNA degradation.

Inactive Publication Date: 2017-02-22
山东艾莱普生物科技有限公司
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Problems solved by technology

The disadvantage is that the silica gel column will adsorb RNA / DNA at the same time, so it is easy to cause DNA residue, resulting in low purity
[0004] The bone tissue is hard, the bone tissue is hard, the bone cell density is low, and the peripheral matrix contains a large amount of mucin (proteoglycan) and RNA is difficult to separate, and high-quality extraction cannot be performed by the traditional Trizol method
In the process of extracting bone tissue RNA, the RNA is often degraded due to long extraction time, RNase contamination and other reasons, making subsequent experiments impossible

Method used

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  • Quick osseous tissue RNA extraction kit

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[0050] The four bone tissues of rat femoral head, rabbit intervertebral disc, rabbit knee articular cartilage, and mouse tibia were operated as follows:

[0051] 1. Put the crushed bone tissue into the mortar after clamping the bone forceps, add liquid nitrogen and grind it into fine powder repeatedly to keep the liquid nitrogen always present. Transfer 100 mg of fine powder to the preheated lysate CLB centrifuge tube. Described lysate contains the guanidine isothiocyanate that concentration is 3mol / L, the hexadecyltrimethylammonium bromide of 2g / L, the Tris-HCl buffer solution that pH is 7.0, the polyvinylpyrrolidone of 2g / L, Sodium chloride with a concentration of 50mmol / L. Immediately vortex vigorously for 45 sec. Place in a 65°C water bath (10 min), and invert twice in the middle to aid lysis.

[0052] 2. Centrifuge the lysate for 10 minutes to precipitate fragments that cannot be lysed. Take the supernatant, add 1 / 2 volume of absolute ethanol to the supernatant, mix we...

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Abstract

The invention discloses a quick osseous tissue RNA extraction kit. The kit comprises at least a genomic DNA removal column, at least one RNA absorptive column, lysis buffer used cooperatively with the genomic DNA removal column and elution buffer used cooperatively with the RNA absorptive column. The kit has the advantages that the exclusive phenol/chloroform-free lysis buffer is used and a plurality of ingredients are added to remove proteoglycan. A particular buffer system and a bimodal pore silica gel purification system are utilized to solve the problem of remaining DNA, thus osseous tissue RNA with high purity and good integrity can be obtained. The particular genomic DNA removal column can effectively remove remaining genomic DNA (gDNA), thus the extracted RNA can be directly used in reverse transcription PCR and real-time PCR without DNase digestion. Extraction of single samples can be completed in 35 minutes generally, and RNA degradation due to long-time extraction, RNase contamination and other causes is avoided.

Description

technical field [0001] The invention relates to RNA extraction, and relates to the field of nucleic acid purification, in particular to a rapid extraction kit for bone tissue RNA. Background technique [0002] The research of molecular biology mainly focuses on nucleic acid and protein. Most molecular biology research needs to obtain purified nucleic acid first. Therefore, nucleic acid purification is one of the cornerstones of molecular biology. The global market is very large, and countries all over the world are doing Continuously improve the extraction and purification methods to achieve faster and purer nucleic acid. Nucleic acid purification is generally divided into two categories: RNA extraction and purification and DNA extraction and purification. [0003] There are mainly two mainstream RNA extraction solutions in the world. One is the guanidine isothiocyanate / acid phenol / chloroform one-step extraction of total RNA (commonly known as the TRIzol method) invented by...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/101
Inventor 张茂
Owner 山东艾莱普生物科技有限公司
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