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Real-time fluorescent RPA kit and test strip RPA kit for rapid detection of porcine type 2 circovirus and application of real-time fluorescent RPA kit and test strip RPA kit

A detection kit and circovirus technology, which are applied in the field of test strip RPA kits and kits for detecting porcine circovirus type 2, to achieve the effects of true and reliable detection results, increased specificity, and reduced product contamination

Active Publication Date: 2016-04-27
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0010] For the ORF2 gene of porcine type 2 circovirus, the present invention establishes and evaluates the RPA (real-timeRPA) detection kit based on fluorescent probe and the LFSRPA detection kit based on test strips to realize rapid detection of porcine type 2 circovirus The purpose of the virus, as far as we know, there is no such kit at home and abroad to detect porcine circovirus type 2

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  • Real-time fluorescent RPA kit and test strip RPA kit for rapid detection of porcine type 2 circovirus and application of real-time fluorescent RPA kit and test strip RPA kit
  • Real-time fluorescent RPA kit and test strip RPA kit for rapid detection of porcine type 2 circovirus and application of real-time fluorescent RPA kit and test strip RPA kit
  • Real-time fluorescent RPA kit and test strip RPA kit for rapid detection of porcine type 2 circovirus and application of real-time fluorescent RPA kit and test strip RPA kit

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Embodiment 1

[0052] Example 1 Rapid detection of porcine type 2 circovirus real-time fluorescent RPA (Real-timeRPA) detection kit and the establishment of detection methods

[0053] 1. Design and preparation of primers and probe sequences

[0054] The inventors of the present invention passed KR559725.1, KR559695.1, KM455975.1, KP768481.1, JN181905.1, JN181902.1, KF850461.1, KC620515.1, KF035059.1, KF951567.1 from GenBank , KF951570.1, KM624031.1 and JQ002672.1 ORF2 gene homologous sequences were compared and analyzed, and the conserved region of porcine circovirus type 2 ORF2 gene was determined, and primers and probes were designed for this region in order to detect to as many porcine circovirus type 2 as possible. All primers and probes were synthesized by Sangon Biotech (Shanghai, China). The present invention designed and synthesized three pairs of upstream primers, three pairs of downstream primers and one probe sequence respectively, as shown in Table 1 below.

[0055] Table 1, p...

Embodiment 2

[0089] Example 2 Establishment of the test strip RPA (LFSRPA) detection kit and detection method for rapid detection of porcine type 2 circovirus

[0090] 1. Design and preparation of primers and probe sequences

[0091] The inventors of the present invention passed KR559725.1, KR559695.1, KM455975.1, KP768481.1, JN181905.1, JN181902.1, KF850461.1, KC620515.1, KF035059.1, KF951567.1 from GenBank , KF951570.1, KM624031.1 and JQ002672.1 ORF2 gene homologous sequences were compared and analyzed, and the conserved region of porcine circovirus type 2 ORF2 gene was determined, and primers and probes were designed for this region in order to detect to as many porcine circovirus type 2 as possible. All primers and probes were synthesized by Sangon Biotech (Shanghai, China). The present invention designed and synthesized three pairs of upstream primers, three pairs of downstream primers and one probe sequence respectively, as shown in Table 4 below.

[0092] Table 4, LFSRPA primers ...

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Abstract

The invention discloses a real-time fluorescent RPA kit for rapid detection of porcine type 2 circovirus and application of test strip RPA kit, and also discloses a test strip RPA kit for rapid detection of the porcine type 2 circovirus and application of the test strip RPA kit. The two kits respectively include primers, as shown in SEQ ID NO.1 and SEQ ID NO.2, as well as corresponding probes, and the primers and the probes, although for a same target sequence, are different in modified base and testing platform. Experiments prove that the sensitivities of both kits are 102 copies / reaction and the kits are capable of achieving the specific detection of the porcine type 2 circovirus. Results of detecting suspected samples of the porcine type 2 circovirus with the two kits show that the detection results of the two kits are completely consistent with that of qPCR. Therefore, the two kits of the invention can be used for rapidly, effectively and sensitively detecting the porcine type 2 circovirus; and an effective technical means is provided for the differentiation and the diagnosis of the porcine type 2 circovirus.

Description

technical field [0001] The invention relates to a kit for detecting porcine type 2 circovirus and its application, in particular to a real-time fluorescent RPA kit and test strip RPA kit for rapidly detecting porcine type 2 circovirus based on RPA reaction , and also relates to the application of the two in rapid detection of porcine circovirus type 2, and the invention belongs to the field of preventive veterinary inspection. Background technique [0002] Thirty years have passed since the birth of PCR technology. From classic PCR, real-time quantitative PCR to the current digital PCR, this technology is constantly changing but has never faded out of our field of vision. However, this technology requires special and expensive thermal cycle instruments and skilled operators, which is time-consuming and labor-intensive, and it is difficult to be used in on-site diagnosis and places with poor experimental conditions. In the laboratories of developing countries, due to the li...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/68C12R1/93
CPCC12Q1/6844C12Q1/701C12Q2521/507C12Q2563/107C12Q2565/625
Inventor 张志东杨洋秦晓东孙英军
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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