Kit and method for extracting nucleic acids from urine

A technology for extracting nucleic acid and extraction method, which is applied in the field of biology, can solve the problems of long sample loading time, loss of free nucleic acid, unfavorable nucleic acid extraction, etc., and achieves the effect of simple operation, high purity of nucleic acid, and saving operation time

Inactive Publication Date: 2014-12-10
宁波有成生物医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, such a sample loading method requires too long sample loading time for urine with low nucleic acid content and large sample volume, which is not conducive to the extraction of nucleic acid.
There is also a method to centrifuge the urine first, and then take the precipitated cells as a sample, which can effectively reduce the volume of the sample, but this kind of extraction method will lose the free nucleic acid in the urine supernatant, and the free nucleic acid in the urine contains a large amount. biological information

Method used

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  • Kit and method for extracting nucleic acids from urine
  • Kit and method for extracting nucleic acids from urine
  • Kit and method for extracting nucleic acids from urine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Take 10mL of fresh urine from a healthy person, add 15mL of lysate ULB, mix by inverting, and bathe in water at 37°C for 10 minutes. Assemble the suction filtration device to the suction filtration bottle to configure a complete suction filtration system (see figure 1 ). Pour the sample into the sample tube, start the air pump, and after the sample completely passes through the adsorption column, turn off the air pump and take out the adsorption column. Add 500 μL of LUWB1 to the adsorption column, centrifuge at 12000 rpm for 1 min, add 500 μL of LUWB2, centrifuge at 12000 rpm for 1 min, and finally use 35 μL of eluent for elution.

[0035] PCR amplification experiment: take human GAPDH as the target gene, take 2 μL of the eluted nucleic acid solution as a PCR template, add 11 μL of deionized water and 7 μL of premix, and the reaction system is 20 μL. The premix preparation method is shown in Table 5, and the PCR reaction program is shown in Table 6.

[0036] Primer ...

Embodiment 2

[0044] Take 20mL of urine from a patient diagnosed with pancreatic cancer, and do two parallel experiments, each tube of urine sample is 10mL. Add 15mL lysate solution ULB to each, invert and mix well, and bathe in water at 37°C for 10 minutes. Assemble the suction filtration device to the suction filtration bottle to configure a complete suction filtration system (see figure 1). Pour the sample into the sample tube, start the air pump, and after the sample completely passes through the adsorption column, turn off the air pump and take out the adsorption column. Add 500 μL of LUWB1 to the adsorption column, centrifuge at 12000 rpm for 1 min, add 500 μL of LUWB2, centrifuge at 12000 rpm for 1 min, and finally use 35 μL of eluent for elution.

[0045] PCR amplification experiment: take the human mutated KRAS as the target gene, take 2 μL of the eluted nucleic acid solution as a PCR template, add 11 μL of deionized water and 7 μL of the premix, the reaction system is 20 μL, and...

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Abstract

The invention relates to a method for extracting nucleic acids with urine as a sample in the biological field, and application thereof, and provides a special kit and method for rapidly extracting the nucleic acids from a large amount of urine. The method comprises the following steps: enriching the nucleic acids in the urine onto a nucleic acid adsorbing column in a suction filtration column manner, washing, eluting and the like for extracting a trace amount of nucleic acids in the urine. The nucleic acids obtained by using the kit provided by the invention comprise free nucleic acids in the urine and nucleic acids in exfoliated cells in the urine; the operation is simple and rapid; the nucleic acids are high in yield and high in purity; the kit and the method can be directly applied to molecular biological detection.

Description

technical field [0001] The invention relates to a method for extracting nucleic acid (DNA and RNA) in the field of biology, in particular to a method for extracting nucleic acid from human urine and its application. Background technique [0002] Urine is formed after blood flows through the kidneys, filtered by the glomeruli, reabsorbed and secreted by the renal tubules, and is excreted through the bladder and urethra. Urine is the final metabolite to keep the internal environment of the body stable. In addition to water, glucose, inorganic salts, protein, urea, and uric acid, it also contains red blood cells, white blood cells, exfoliated epidermal cells, free DNA and RNA (mRNA, miRNA) Wait). The free nucleic acid in the urine includes the free nucleic acid from the systemic circulation filtered by the kidney and the nucleic acid released from the cell lysis of the urinary tract. Urine contains a wealth of biological information, and has the advantages of non-invasive col...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
Inventor 唐涛周细武邱英华
Owner 宁波有成生物医药科技有限公司
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