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Detection method of enteromorpha micro-generation fluorescence in-situ hybridization

A fluorescent in situ hybridization and detection method technology, applied in the field of green tide biological detection and monitoring research, can solve the problem of less research, achieve the effect of short time, good specificity, and time saving

Inactive Publication Date: 2010-10-06
YELLOW SEA FISHERIES RES INST CHINESE ACAD OF FISHERIES SCI
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Problems solved by technology

So far, molecular probe technology has developed rapidly in the research on red tide biological detection and monitoring in the world, but there are very few related studies on green tide, especially on the microgeneration of Enteromorpha prolifera, that is, the overwintering seed bank ( Fluorescent in situ hybridization in the form of propagule bank) has not been reported

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  • Detection method of enteromorpha micro-generation fluorescence in-situ hybridization
  • Detection method of enteromorpha micro-generation fluorescence in-situ hybridization
  • Detection method of enteromorpha micro-generation fluorescence in-situ hybridization

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Embodiment

[0072] 1. Obtaining samples

[0073] The experiment started in June 2008 and ended in July 2008. The water body samples were collected from the intertidal zone with dense floating Enteromorpha in Qingdao Second Beach (35.35°N, 119.30°E, 10-35cm below the water surface) on June 15, 2008. The operation process after collecting the samples is the same as the above-mentioned experimental operation. A total of 55 water samples were collected, each 5L. In the laboratory, use a 10μm sieve to filter and concentrate to reduce the volume of water to 100mL.

[0074] 2. Probe design

[0075] The oligonucleotide probes used in Enteromorpha for FITC-labeled in situ hybridization (designed according to the ITS sequence) were designed in the same way as the above method, and the 5' end was labeled with FITC. A total of 4 probes were designed:

[0076] 1) 5'-GCCCGCCGTTTACAGGAT-3'

[0077] 2) 5'-AGCCCTAACCCATTGAACCC-3'

[0078] 3) 5'-CCCTGCGATAGTAACTGAGACA-3'

[0079] 4) 5'-GGAGCCCTAACCC...

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Abstract

The invention relates to green-tide biological detection and monitoring research, in particular to a detection method of enteromorpha micro-generation fluorescence in-situ hybridization. An oligonucleotide probe for marking the in-situ hybridization of enteromorpha micro-generation with FITC (Fluoresceine Isothiocyanate) comprises one or more than one of the following four probes: (1) 5'-gcccgccgtttacaggat-3', (2) 5'-agccctaacccattgaaccc-3', (3) 5'-ccctgcgatagtaactgagaca-3' and (4) 5'-GGAGCCCTAACCCATTGAA-3', wherein the 5' terminals are marked by an FITC marking method. By the invention, the enteromorpha micro-generation can be rapidly and accurately evaluated and counted to confirm the biomass and the distribution situation thereof and achieve the aim of accurately detecting and forecasting.

Description

technical field [0001] The invention relates to green tide biological detection and monitoring research, in particular to a fluorescence in situ hybridization detection method of enteromorpha microgeneration. Background technique [0002] Detection methods based on molecular basis (nucleic acid, protein, glycoprotein) have great advantages over traditional detection methods, species identification is more accurate and reliable, and can provide quantitative information, and can detect low-abundance target microalgae species (large micro-generations of algae), enabling their use in routine monitoring. Due to the heterogeneity of the rRNA gene ITS region, it shows obvious differences among various genera and species. Internationally, the ITS area is usually used as an area for the classification and systematic research of various biological groups and genera, and it is also the first choice for designing probes. Relatively speaking, the ITS region, the transcriptional spacer ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G01N21/64C12N15/11
Inventor 张晓雯叶乃好毛玉泽庄志猛王清印
Owner YELLOW SEA FISHERIES RES INST CHINESE ACAD OF FISHERIES SCI
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