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Kit capable of detecting natural water sample of prorocentrum donghaiense in high throughput manner

A technology of Prorocentrum and kit, applied in the field of kits capable of high-throughput detection of natural water samples of Prorocentrum in the East China Sea

Inactive Publication Date: 2013-05-08
HARBIN INST OF TECH AT WEIHAI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] It can be seen that there is currently no report on a high-throughput detection technology for red tide algae water samples and its corresponding kits

Method used

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  • Kit capable of detecting natural water sample of prorocentrum donghaiense in high throughput manner

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Embodiment Construction

[0026] The present invention will be described in detail below in conjunction with specific embodiments.

[0027] 1. Kit composition

[0028] (1) Reagent A is a crude nucleic acid extraction reagent for natural samples, and its composition is: 2% (m / v) CTAB, 0.5% (v / v) β-mercaptoethanol, 1.4 M NaCl, 20 mM EDTA, 100 mM Tris-Cl , 10 U RNase Inhibitor;

[0029] (2) Reagent B is 5×NASBA Buffer, its composition is 0.2 M Tris-HCl, 60 mM MgCl 2 , 0.35 M KCl 2 , 25 mM DTT, 20 mM dNTP;

[0030] (3) Reagent C is NTP (10 mM each);

[0031] (4) Reagent D is 5×Primer mix, its composition is: 75% DMSO, 1 μM NASBA-P1, 1 μM NASBA-P2, 25 mM DTT, 20 mM dNTP;

[0032] (5) Reagent E is 4×Enzyme mix, its composition is: 1.5 M sorbitol, 0.42 mg / ml BSA, 16 U / ml RNase H, 6400 U / ml T7 RNA Polymerase, 1280 U / ml RT-M-MLV , 4000 U / ml RNA inhibitor;

[0033] (6) Reagent F is RNA denaturant, its composition is: 1.5 M NaCl, 0.15 M trisodium citrate dihydrate, pH 7.0, 0.15 M formaldehyde;

[0034] (...

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Abstract

The invention discloses a kit capable of detecting a natural water sample of prorocentrum donghaiense in a high-throughput manner. The kit comprises 12 constituents, including a natural sample nucleic acid crude-extract reagent, 5*NASBA (Nucleic Acid Sequence Based Amplification) Buffer, 5*Primer mix, 4*Enzyme mix, an RNA (Ribonucleic Acid) denaturing agent and a hybridization solution. The detection principle of the kit is as follows: rRNA (Ribosomal Robonucleic Acid) of an algae cell is taken as a target sequence which is prepared by using an isothermal nucleic acid amplification technology, and a plurality of detection samples are detected at the same time by virtue of the reverse dot blot of RNA. According to the kit, a detection process is in need of simple equipment only, the detection is rapid, a detection result can be judged directly through naked eyes, and the kit has high sensitivity, can be used for site detection of a daily water environment and has a certain practical value for early warning of the red tide of the prorocentrum donghaiense.

Description

technical field [0001] The invention relates to the technical field of marine biology, in particular to a kit for high-throughput detection of natural water samples of Prorocentrum donghaiense. Background technique [0002] Red tides are the discoloration of sea water due to the overgrowth or accumulation of marine plankton, including algae (the main species), protozoa and bacteria. In recent years, due to the intensification of human activities, the ocean has become increasingly eutrophic, resulting in frequent red tides, which have become a global ecological disaster, causing huge losses to the aquaculture industry, fishing industry and coastal tourism. The marine ecological environment has a negative impact and even directly threatens human health. [0003] The correct identification of the causative species of red tides (including Prorocentrum donghaiense) is the premise and basis for the study, understanding and control of red tides caused by them. At the same time, si...

Claims

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Application Information

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IPC IPC(8): G01N21/78
Inventor 陈国福张春云
Owner HARBIN INST OF TECH AT WEIHAI
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