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Specific molecular marker primers, kit and identification method for Thymallus arcticus grubei population

An identification method and molecular marker technology, applied in biochemical equipment and methods, microbe determination/inspection, DNA/RNA fragments, etc., can solve the problem of depletion of wild resources, change of spawning, feeding, overwintering environment of Heilongjiang grayling, inability to Issues such as the amount of catch

Active Publication Date: 2019-07-23
HEILONGJIANG RIVER FISHERY RES INST CHINESE ACADEMY OF FISHERIES SCI
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  • Abstract
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  • Application Information

AI Technical Summary

Problems solved by technology

This fish has strict requirements on environmental conditions, and it is difficult to form a large-scale group in a natural state, let alone a certain amount of fishing
At the same time, due to changes in environmental conditions, fishermen in habitats fished during the migration period and built reservoirs and water conservancy facilities, which changed the spawning, feeding, and overwintering environments of Heilongjiang grayling, which had a great impact on its reproduction and survival, resulting in the loss of wild resources. exhausted

Method used

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  • Specific molecular marker primers, kit and identification method for Thymallus arcticus grubei population
  • Specific molecular marker primers, kit and identification method for Thymallus arcticus grubei population
  • Specific molecular marker primers, kit and identification method for Thymallus arcticus grubei population

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Embodiment 1

[0030] 1. Primer design

[0031] The RNA of Heilongjiang grayling was sequenced to obtain the EST library of Heilongjiang grayling, and the microsatellite loci with 2, 3, 4, 5 repeat sequences were searched in the EST library. 50 sequences were randomly selected, and Primer 3 was used to design primers with a length of 19–25 bp and a GC content of 50–60%. Screening of 50 pairs of primers revealed that ThtC1010, ThtC1029 and ThtC1034 markers had population specificity.

[0032] Primer sequences for population-specific molecular markers of Heilongjiang grayling:

[0033] ThtC1010 upstream primer: 5'-TCAGGTTGTGCAAAATCTCGA-3' (SEQ ID NO.1)

[0034] Downstream primer: 5'-GGCCAGCAGAACCAGAATCA-3' (SEQ ID NO.2)

[0035] ThtC1029 upstream primer: 5'-GTGTTTCTTCCTGGCAGGGA-3' (SEQ ID NO.3)

[0036] Downstream primer: 5'-TGGTGGTTAAGACTCAGAGACC-3' (SEQ ID NO.4)

[0037] ThtC1034 upstream primer: 5'-TGGTGACACTGAAGAGGGGA-3' (SEQ ID NO.5)

[0038] Downstream primer: 5'-AGCGTGTTATTACCAGTG...

Embodiment 2

[0052] The fin ray samples of 20 tails of Genhe grayling, 20 tails of Taoer river grayling and 26 tails of Wusuli river grayling were collected, and identified by the identification method in Example 1. According to the comparison between the electrophoresis results and the provided standard map, it was found that the primer ThtC1010 amplified the band size of 20 graylings in the Genhe population with the same 109bp as A, and the amplified bands of 20 graylings in the Taoerhe population and 26 graylings in the Wusulijiang population were amplified. The size of the enlarged band is the same as B, 114bp; primer ThtC1029, the amplified band size of 20 graylings in Genhe population is the same as C, 198bp, and the amplified bands of 20 graylings in Taoerhe population and 26 graylings in Wusulijiang population The band size is the same as D 184bp; primer ThtC1034 Genhe population amplified band size is the same as E, F and G respectively 174bp, 168bp and 164bp, of which 5 tails have...

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Abstract

The invention discloses specific molecular marker primers, a kit and an identification method for Thymallus arcticus grubei population. The primers are ThtC1010, ThtC1029 and ThtC1034, the sequence ofthe primer ThtC 1010 is shown in SEQ ID NO. 1-2, the sequence of the primer ThtC1029 is shown in SEQ ID NO. 3-4, and the sequence of the primer ThtC 1034 is shown in SEQ ID NO. 5-6. According to thepresent invention, the method is a new method based on the principle of molecular biology technology for distinguishing the Thymallus arcticus grubei population in China and other countries for the first time. EST-SSR markers are successfully developed on the basis of EST data of Thymallus arcticus grubei, the genetic background of the Thymallus arcticus grubei population is analyzed with partialmarkers, and specific markers capable of distinguishing three populations are found. Genhe graylling population can be accurately distinguished from the other two populations by a THC1010 locus and aTHC1029 locus, the three populations can be distinguished by a THC1034 locus, and the method has the advantages of high accuracy, good technical reproducibility and high cost performance.

Description

technical field [0001] The invention relates to a Heilongjiang grayling population-specific molecular marker primer, a kit and an identification method, belonging to the technical field of molecular biology. Background technique [0002] Heilongjiang grayling Thymallus arcticus grbei (Dybowski) belongs to Salmoniformes (Salmoniformes), grayling family (Thymallidae), grayling genus (Thymallus), commonly known as: spotted trout, red scale fish, distributed in Eurasia, and is naturally found in my country. Distributed in the tributaries of the Heilongjiang River Basin, the meat is tender and delicious, and it is a rare and precious cold-water fish unique to the Heilongjiang River System. This fish has strict requirements on environmental conditions, and it is difficult to form a large-scale group in a natural state, let alone a certain amount of fishing. At the same time, due to changes in environmental conditions, fishermen in habitats fished during the migration period and bu...

Claims

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Application Information

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IPC IPC(8): C12Q1/6888C12N15/11
CPCC12Q1/6888C12Q2600/156Y02A40/81
Inventor 佟广香唐国盘董乐杨笑星匡友谊尹家胜
Owner HEILONGJIANG RIVER FISHERY RES INST CHINESE ACADEMY OF FISHERIES SCI
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