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Method for extracting total RNA in mid-intestine of saperda populnea

The invention relates to a technology for the extraction method of A. chinensis, which is applied in chemical instruments and methods, preparation of sugar derivatives, sugar derivatives, etc., and can solve the problems of low yield, complicated operation and high cost of extracting RNA, and achieves fast and simple operation, high yield, high cost and high cost. Yield full effect

Inactive Publication Date: 2009-09-23
NORTHEAST FORESTRY UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] The purpose of the present invention is to solve the problems of complex operation, time-consuming, high cost, low yield of extracted RNA, and easy degradation in existing methods for extracting insect midgut RNA, and to provide a method for extracting total RNA from the midgut of Populus chinensis

Method used

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  • Method for extracting total RNA in mid-intestine of saperda populnea
  • Method for extracting total RNA in mid-intestine of saperda populnea

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specific Embodiment approach 1

[0007] Embodiment 1: In this embodiment, the method for extracting total RNA from the midgut of Populus chinensis is realized according to the following steps: 1. Add 500-700 μL of 2×CTAB buffer solution and 0.5-1.0 mg of polyvinylpolypyrrolidone to 1.5 mL of Mix well in the Eppendorf centrifuge tube, and in the liquid nitrogen environment, grind the midgut of Cyperus chinensis and add it to the Eppendorf centrifuge tube, and mix well; 2. Put the Eppendorf centrifuge tube in a water bath at a temperature of 55-65°C for 10 minutes; 3. Add 500 μL ~ 700 μL of chloroform / isoamyl alcohol into an Eppendorf centrifuge tube, and centrifuge for 5 minutes at a centrifugal speed of 10,000 ~ 13,000 r / min; 4. Take the centrifuged supernatant and put it into a new Eppendorf centrifuge tube, Repeat step 3; 5. Take the supernatant after centrifugation in step 4 and put it into another new Eppendorf centrifuge tube, then add LiCl solution with a volume of 500-700 μL and a molar concentration of...

specific Embodiment approach 2

[0011] Specific embodiment two: the difference between this embodiment and specific embodiment one is that the concentration of Tris-HCl in the 2×CTAB buffer solution in step one is 100mmol / L, the concentration of EDTA is 20mmol / L, and the concentration of NaCl is 1.4mol / L , the mass fraction of CTAB is 2%-3%, the volume fraction of β-mercaptoethanol is 0.4%, and the mass fraction of polyvinylpolypyrrolidone is 1%. Other steps and parameters are the same as those in Embodiment 1.

specific Embodiment approach 3

[0012] Specific embodiment three: the difference between this embodiment and specific embodiment one is that the concentration of Tris-HCl in the 2×CTAB buffer solution in step one is 100mmol / L, EDTA is 20mmol / L, and the concentration of NaCl is 1.4mol / L. L. The mass fraction of CTAB is 2%, the volume fraction of β-mercaptoethanol is 0.4%, and the mass fraction of polyvinylpolypyrrolidone is 1%. Other steps and parameters are the same as those in Embodiment 1.

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Abstract

The invention discloses a method for extracting total RNA in the mid-intestine of saperda populnea, relating to the extraction method for the RNA in the mid-intestine of insects. The extraction method solves the problems that the existing extraction method for the RNA in the mid-intestine of insects has complex operation, high cost, low RNA extraction output and easy degradation, and takes time. The method comprises the following steps: firstly, 2*CTAB buffer solution and crospovidone are added into a centrifugal tube, and then the mid-intestine of saperda populnea ground by a liquid nitrogen is added in the centrifugal tube to be well blended; secondly, the centrifugal tube is processed by hot bath; thirdly, chloroform / isoamylol is added into the centrifugal tube to be centrifugated; fourthly, a supernatant fluid is taken out, and the step three is repeated; fifthly, the supernatant fluid centrifugated in the step four is added with LiCl solution, cooled, stands and then centrifugated; sixthly, the supernatant fluid is discarded, washed by alcohol, and centrifugated; seventhly, after the supernatant fluid in the step six is discarded, the alcohol is evaporated, and then the sterile water is added. The method has brief operation, low cost, high success ratio, saves time and can obtain complete insect RNA with high purity, higher output and difficult degradation.

Description

technical field [0001] The invention relates to a method for extracting insect midgut RNA. Background technique [0002] Nucleic acid extraction is the basis of molecular biology experiments, and high-quality nucleic acid is a necessary prerequisite for downstream technologies such as molecular markers, gene cloning, and gene expression research. However, at present, the methods for extracting insect midgut RNA mainly include guanidine isothiocyanate method and Trizol method, but these methods have the problems of complicated operation, time-consuming, high cost, low yield of extracted RNA, and easy degradation. Contents of the invention [0003] The purpose of the present invention is to solve the problems of complex operation, time-consuming, high cost, low yield of extracted RNA and easy degradation in the existing insect midgut RNA extraction method, and to provide a method for extracting total RNA from the midgut of Populus chinensis. [0004] The method for extracti...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07H21/02C07H1/08
Inventor 迟德富姚大彬关桦楠李晓灿宇佳
Owner NORTHEAST FORESTRY UNIVERSITY