Fluorescent probe PCR (polymerase chain reaction) method for simultaneously detecting Edwardsiella tarda and Edwardsiella ictaluri

Abstract

The invention relates to a microbe detection method and detection kit, particularly a fluorescent probe PCR (polymerase chain reaction) method for simultaneously detecting Edwardsiella tarda and Edwardsiella ictaluri. The method comprises the following step: carrying out real-time fluorescence PCR reaction by using bacteria DNA (deoxyribonucleic acid) as a template by utilizing specific probes and primer pairs. The method is characterized in that the primer primers are respectively sequences disclosed as SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3 and SEQ ID NO.4; the SEQ ID NO.1 is SEQ ID NO.1: 5'-CTGCCAAGCAGGGACGTAA-3', SEQ ID NO.2: 5'-CGAGGAGAGCATCTTGTCGAA-3', SEQ ID NO.3: 5'-TCCATTCGTCTGTGCGACAA-3', and SEQ ID NO.4: 5'-AAAACACGTTCGGATGGATTG-3'; the probes are nucleotide sequences which is identical to or complementary with the sequences between the primer pair on the Edwardsiella tarda or Edwardsiella ictaluri genome DNA; the 5' end and 3' end of the probes are respectively labeled with a fluorescence report group and a fluorescent quenching group; the probes are sequences disclosed as SEQ ID NO.5 and SEQ ID NO.6; and the SEQ ID NO.5 is 5'-ATCCTCAACGTCGAGAAGGCGCG-3', and the SEQ ID NO.6 is 5'-CACTATGAGGGTGGGATCAAGGCGTTT-3'.

Description

technical field [0001] The invention relates to a detection method and a detection kit for microorganisms, in particular to primers and probe sequences and a kit for simultaneous detection of Edwardsiella tarda and Edwardsiella spp. for real-time fluorescent PCR detection. Background technique [0002] Edwardsiella tarda ( Edward Siella sp.) is widely distributed in seawater and freshwater, infects various farmed aquatic animals, and is extremely harmful to aquaculture. It is pathogenic to eels, catfish, squid and other animals, and can cause serious economic losses. The pathogenic bacteria that cause fish disease are mainly Edwardsiella tarda ( E. tarda ) and Edwardsiella ( E. ictaluri ). It is now known that Edwardsiella tarda is an important pathogenic bacterium of a variety of freshwater and seawater cultured fish. It causes diseases of fish kidney and liver abscess lesions, and the loss is serious. It is a pathogenic bacterium that is more harmful in aquaculture a...

AI Technical Summary

Problems solved by technology

[0003] At present, the Edwardsiella gene detection technology established at home and abroad includes the common PCR detection technology for detecting Edwardsiella tarda, and the common PCR detection technology for detecting Edwardsiella spp., these methods need many times to detect these two bacteria, and Gel electrophoresis is required to observe the test results, and the detection method is more cumbersome and time-consuming than the fluorescent PCR method

[0004] At present, there is no fluorescent probe PCR detection method for the simultaneous detection of Edwardsiella tarda and Edwardsiella spp. at home and abroad.

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