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Method for obtaining genome sequence of papaya ringspot virus through high-throughput small RNA sequencing

A virus genome and ringspot virus technology, applied in the fields of virus peptide, genetic engineering, DNA preparation, etc., can solve the problems of economic loss, papaya production and harvest, etc.

Inactive Publication Date: 2013-01-30
INST OF TROPICAL BIOSCI & BIOTECH CHINESE ACADEMY OF TROPICAL AGRI SCI
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  • Abstract
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  • Application Information

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Problems solved by technology

At the same time, papaya virus disease has become the biggest limiting factor restricting the development of papaya industry, and in some areas even led to the extinction of papaya production and harvest, resulting in huge economic losses

Method used

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  • Method for obtaining genome sequence of papaya ringspot virus through high-throughput small RNA sequencing
  • Method for obtaining genome sequence of papaya ringspot virus through high-throughput small RNA sequencing
  • Method for obtaining genome sequence of papaya ringspot virus through high-throughput small RNA sequencing

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Embodiment 1

[0062] A method for cloning the whole genome sequence of papaya ringspot virus Hainan isolate PRSV-HN-y1 obtained by high-throughput Small RNA sequencing, comprising the following steps:

[0063] (1) Total RNA was extracted from samples infected with papaya mosaic disease, sRNA was separated by polyacrylamide gel electrophoresis (PAGE gel), the sequence information was trimmed after Small RNA sequencing, and the linker sequence was removed to obtain sRNA fragments.

[0064] (2) Compare the sRNA fragments with all known viral genome sequences in Genbank by running Bowtie software (default parameters), and screen out highly matching small RNA fragments; as attached figure 1 shown.

[0065] (3) Run the Velvet software, set the parameters to k-mer=17 and minimum coverage=5, use the small RNA fragments obtained in step 2 as input, and assemble them into short contig sequences; as attached figure 2 , 3 , 4 shown.

[0066] (4) Run the Blast software (default parameters) to compa...

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Abstract

The invention belongs to the fields of bioinformatics and genetic engineering. A papaya ringspot virus Hainan isolate PRSV-HN-y1 has genome sequences of SEQ ID No. 1 and SEQ ID No. 2 as shown in the specification. The invention further discloses a cloning method for obtaining the genome sequence of the papaya ringspot virus Hainan isolate PRSV-HN-y1 through a high-throughput small RNA sequencing. Research results firstly confirm that the high-throughput small RNA sequencing can be used for pathogen identification, detection and virus whole genome sequencing of samples infected by suspected papaya virus.

Description

technical field [0001] The invention belongs to information comparison, splicing, assembly and molecular cloning technologies in the fields of bioinformatics and genetic engineering, and specifically relates to a method for obtaining the genome sequence of papaya virus by high-throughput Small RNA sequencing, which is used for the discovery and function of new viruses. Research provides strong technical support. Background technique [0002] High-throughput sequencing technology has been widely used in the genome, transcriptome, gene expression regulation research of eukaryotes, prokaryotes, viruses, etc. The specific analysis techniques include: 1De novo re-sequencing, 2DNA re-sequencing, 3RNA transcriptome, expression Spectrum sequencing, 4MicroRNA discovery and analysis, 5methylation sequencing, 6DNA-protein interaction research, 7target region capture sequencing, 8exon sequencing. At present, Illumina has recently launched a new sequencer Hiseq2000, which is mainly used...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/40C07K14/08C12N15/10G06F19/22
Inventor 张雨良刘志昕熊忠国黄启星史学群王健华余乃通章绍延
Owner INST OF TROPICAL BIOSCI & BIOTECH CHINESE ACADEMY OF TROPICAL AGRI SCI
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