Primer and method for detecting leuciscus waleckii in Dalai Nur lake basin

A technique for detecting primers for crocodile vaschii, applied in biochemical equipment and methods, measurement/inspection of microorganisms, DNA/RNA fragments, etc., can solve problems such as difficulties in protecting unique wild fishery resources, achieve high accuracy, The effect of simple detection steps and strong operability

Inactive Publication Date: 2014-01-08
HEILONGJIANG RIVER FISHERY RES INST CHINESE ACADEMY OF FISHERIES SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Moreover, it has also brought difficulties to the protection of unique wild fishery resources in our country.

Method used

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  • Primer and method for detecting leuciscus waleckii in Dalai Nur lake basin
  • Primer and method for detecting leuciscus waleckii in Dalai Nur lake basin

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specific Embodiment approach 1

[0019] Specific embodiment 1: In this embodiment, the primers for the detection of crocodile in the Dali Lake Basin are a pair of microsatellite marker primers, the nucleotide sequence of the forward detection primer is 5'-ACTTTACATTGGTGCTA-3', and the reverse detection primer nucleotide sequence The acid sequence is 5'-GATTTGGTCAAAGAAGA-3'.

specific Embodiment approach 2

[0020] Specific embodiment two: the present embodiment Dali Lake Basin Washi Yaluo fish detection method is carried out according to the following steps:

[0021] 1. Sample fish DNA extraction:

[0022] Take the fin samples and place them in a 1.5ml centrifuge tube, then add 600μl lysate and 10μl proteinase K in sequence, mix well and place the centrifuge tube in a water bath at 55°C for overnight digestion; add 10μl RNase after digestion ℃ water bath in a water bath for 1 hour; then add 600 μl of a mixture of phenol and chloroform, mix well, and then centrifuge at 12,000 r / min at room temperature for 10 minutes; take the centrifuged supernatant and put it in a new centrifuge tube, then add 1ml of absolute alcohol and mix well, Centrifuge at 12000r / min for 10min; spin to pour out the alcohol after centrifugation, then add 500μl of 70% ethanol, centrifuge at 12000r / min for 5min at room temperature; then remove the ethanol with a pipette gun, and centrifuge the white DNA precipi...

specific Embodiment approach 3

[0031]Specific embodiment 3: The difference between this embodiment and specific embodiment 2 is that the 148bp band amplified in step 3 is subjected to polyacrylamide gel electrophoresis, and then recovered, purified and transferred into competent cells E.coli DH5α , and then extract the plasmid for sequencing to obtain the base sequence as shown in SEQ ID NO:3. Other steps and parameters are the same as in the second embodiment.

[0032] Recover the 148bp strip into a 1.5ml centrifuge tube, then add 50 μl of sterile water, and then centrifuge at 12,000 r / min for 10 to 15 minutes in a boiling water bath for 15 to 30 minutes (the DNA will be precipitated from the strip at this time). Using the recovered product as a template, PCR amplification is performed again with the detection primers in the first embodiment, and the amplification system and amplification conditions are the same as those in the second embodiment. Carry out carrier ligation reaction after PCR amplification...

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Abstract

The invention discloses a primer and method for detecting leuciscus waleckii in the Dalai Nur lake basin and relates to a primer and method for detecting the leuciscus waleckii. The primer for detecting the leuciscus waleckii in the Dalai Nur lake basin is a pair of microsatellite marker primers. A forward primer nucleotide detection sequence is shown as SEQ ID NO:1 and a backward primer nucleotide detection sequence is shown as SEQ ID NO:2. The method for detecting the leuciscus waleckii in the Dalai Nur lake basin comprises the steps that 1, the DNA of a sample is extracted; 2, PCR amplification is conducted; 3, gel electrophoresis is conducted on the capillary tubes, a sample fish with a 148bp stripe amplified is the leuciscus waleckii in the Dalai Nur lake basin, and if not, the sample fish is not the leuciscus waleckii in the Dalai Nur lake basin. The primer and method can be applied to the fields of fishery detection and animal resource protection.

Description

technical field [0001] The invention relates to a detection primer and a detection method of crocodile. Background technique [0002] Leuciscus waleckii (Leuciscus waleckii), also known as Northeast Leuciscus waleckii, is commonly known as Huazi fish, slippery fish, and white fish. Small cold water economic fish. The total length of the crocodile is about 40 cm, the body length is flat, the abdomen is round, without ventral ribs, the snout is blunt and slightly raised; the mouth is at the end, the upper jaw is slightly longer than the lower jaw, and the upper jaw extends to the lower edge of the eye; the lip is thin, No horny margins, no beards; large eyes, medium-sized scales, complete lateral line, slightly curved ventrally, extending back to the middle axis of the caudal peduncle; no hard spines on the dorsal fin; the back of the body is grayish brown, and the abdomen is silvery white; the base of the scales has obvious Radial lines, gray trailing edge; fins off-white, ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6888C12Q2600/156
Inventor 常玉梅梁利群唐然孙效文窦新杰
Owner HEILONGJIANG RIVER FISHERY RES INST CHINESE ACADEMY OF FISHERIES SCI
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