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Extractant for extracting RNA (Ribose Nucleic Acid) of mulberry leaf and application of extractant

A technology of extractant and leaves, which is applied in the field of extractant for extracting RNA from mulberry leaves, can solve the problems of high cost and cumbersome steps, and achieve the effect of low cost, simple operation steps and high yield

Inactive Publication Date: 2015-04-29
BIOMARKER TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But its disadvantage is that it needs an additional DNA removal step, which is cumbersome and expensive

Method used

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  • Extractant for extracting RNA (Ribose Nucleic Acid) of mulberry leaf and application of extractant
  • Extractant for extracting RNA (Ribose Nucleic Acid) of mulberry leaf and application of extractant
  • Extractant for extracting RNA (Ribose Nucleic Acid) of mulberry leaf and application of extractant

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] A method for using mulberry leaf RNA extractant, comprising the following steps:

[0036] Step a) processing the sample: weigh fresh or ultra-low temperature frozen mulberry leaves, take 1 g, mix it with 0.04 g of polyvinylpyrrolidone, and grind it into powder in liquid nitrogen.

[0037] Step b) preparation of extractant: add 0.1mol Tris-HCl, 0.5mol NaCl, 10mmol EDTA, 20g SDS, 1mL RNase solid phase scavenger to 1L double distilled water. Configure in a volumetric flask and let it stand at room temperature for 24 hours. Wherein the final concentration of Tris-HCl is 0.1mol / L, the final concentration of NaCl is 0.5mol / L, the final concentration of EDTA is 0.01mol / L, the mass volume ratio of SDS is 2%; the volume ratio of solid phase scavenger is 0.1% .

[0038] Step c) extraction: Pour about 100 mg of the powder sample in step a) into a centrifuge tube filled with 1 mL of extractant (4% mercaptoethanol is added before use), and vortex to mix. Centrifuge at 12000 rpm a...

Embodiment 2

[0045] A method for using mulberry leaf RNA extractant, comprising the following steps:

[0046] Step a) processing the sample: weighing ultra-low temperature frozen mulberry leaves, taking 1 g, mixing it with 0.04 g of polyvinylpyrrolidone, and grinding it into powder in liquid nitrogen.

[0047] Step b) preparation of extractant: add 0.105mol Tris-HCl, 0.52mol NaCl, 10.4mmol EDTA, 20.8g SDS, 1.5mL RNase solid phase scavenger to 1L double distilled water. Configure in a volumetric flask and let it stand at room temperature for 24 hours. Wherein the final concentration of Tris-HCl is 0.105mol / L, the final concentration of NaCl is 0.52mol / L, the final concentration of EDTA is 10.4mmol / L, the mass volume ratio of SDS is 2.08%; the volume of solid phase scavenger is 0.15% .

[0048] Step c) extraction: Pour about 100 mg of the powder sample in step a) into a centrifuge tube filled with 1 mL of extractant (4% mercaptoethanol is added before use), and vortex to mix. Centrifuge a...

Embodiment 3

[0055] A method for using mulberry leaf RNA extractant, comprising the following steps:

[0056] Step a) processing the sample: weighing ultra-low temperature frozen mulberry leaves, taking 1 g, mixing it with 0.04 g of polyvinylpyrrolidone, and grinding it into powder in liquid nitrogen.

[0057] Step b) preparation of extractant: add 0.15mol Tris-HCl, 0.75mol NaCl, 15mmol EDTA, 30g SDS, 2mL RNase solid-phase scavenger to 1L double distilled water. Configure in a volumetric flask and let it stand at room temperature for 24 hours. Wherein the final concentration of Tris-HCl is 0.15mol / L, the final concentration of NaCl is 0.75mol / L, the final concentration of EDTA is 15mmol / L, the mass volume ratio of SDS is 3%; the volume ratio of solid phase scavenger is 0.2%.

[0058] Step c) extraction: Pour about 100 mg of the powder sample in step a) into a centrifuge tube filled with 1 mL of extractant (4% mercaptoethanol is added before use), and vortex to mix. Centrifuge at 12000 rp...

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Abstract

The invention provides an extractant for extracting RNA (Ribose Nucleic Acid) of a mulberry leaf. The extractant is prepared by dissolving Tris-HCl, NaCl, EDTA (Ethylene Diamine Tetraacetic Acid), SDS (Sodium Dodecyl Sulfate) and mercaptoethanol into aqueous solution of an RNAse solid phase scavenger, wherein the final concentration of Tris-HCl is 0.05 to 0.2mol / L; the final concentration of NaCl is 0.25 to 1.0mol / L; the final concentration of EDTA is 0.01 to 0.04mol / L; the final concentration of SDS is 1 percent to 4 percent; the final concentration of mercaptoethanol is 2 percent to 8 percent; the mass fraction of the aqueous solution of the RNAse solid phase scavenger is 0.1 percent to 0.2 percent. The extractant provided by the invention is suitable for extracting the RNA of the mulberry leaf, adopts simple operation steps, is low in cost, is high in stability, is high in yield of the extracted RNA and is high in purity of the extracted RNA.

Description

technical field [0001] The invention relates to the technical field of plant genetic engineering, in particular to an extractant for extracting RNA from mulberry leaves and an application thereof. Background technique [0002] Separating and obtaining high-purity RNA is the basis and key to the research of RNA-related technologies. Mulberry leaves are rich in polyphenols and polysaccharides, which will interfere with RNA extraction during the isolation process. Polyphenols are easy to oxidize, and the complexes formed after oxidation (such as quinones) can be stably combined with RNA, resulting in the loss of RNA during the extraction process, thereby affecting the separation and purification of RNA. On the one hand, polysaccharides form jelly and affect the dissolution of RNA; on the other hand, the inhibition of some enzymes affects the application of extracted RNA. The high stability of RNase is also an important reason for the easy degradation of RNA and the failure of...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
Inventor 郑洪坤
Owner BIOMARKER TECH