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Ostertagia circumcincta/marshallagia circumcincta COI gene amplification primer and application thereof

A technology for amplifying primers and nematodes, which is applied in the field of molecular biology, can solve the problems that hinder the application of DNA barcode technology, difficulty in morphological identification, and time-consuming, etc., and achieve the effect of reliable molecular technology guidance

Active Publication Date: 2016-04-20
CHINESE ACAD OF INSPECTION & QUARANTINE
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  • Abstract
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AI Technical Summary

Problems solved by technology

However, relying on morphology to identify nematode species is prone to problems: first, the morphological characteristics of many animal nematode samples are destroyed when specimens are obtained or stored; second, it takes a lot of time to fully grasp the characteristics and differentiation methods of all nematode species, and It is very difficult; thirdly, the morphological characteristics of some different species of ruminant A. / The morphological characteristics of different species of Marshall's nematodes are mainly reflected in the males, and the morphological differences between the females are small, and it is difficult to identify based on the morphology
[0005] In actual research, using the COI universal primers LCO1490 and HCO2198 designed by Folmer et al. in 1994 has a low amplification efficiency in the ruminant Osteria / Marshallia group, which hinders the application of DNA barcoding technology in the above

Method used

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  • Ostertagia circumcincta/marshallagia circumcincta COI gene amplification primer and application thereof
  • Ostertagia circumcincta/marshallagia circumcincta COI gene amplification primer and application thereof
  • Ostertagia circumcincta/marshallagia circumcincta COI gene amplification primer and application thereof

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Embodiment 1

[0026] 1. Sample collection: From 2011 to 2014, a total of 111 individuals of 2 genera and 11 species of ruminant Austeria nematode / Marshall nematode were collected from grassland and pastoral areas in Xinjiang, Inner Mongolia and Qinghai, China.

[0027] 2. DNA extraction: Use phenol-chloroform method or genome extraction kit to extract DNA from collected samples.

[0028] 3. COI universal primer amplification: use the COI universal amplification primer designed by Folmer in 1994

[0029] LCO1490F: 5'-GGTCAACAAATCATAAAGATATTGG-3'

[0030] HCO2198R: 5'-TAAACTTCAGGGTGACCAAAAAATCA-3'

[0031] 4. Perform PCR amplification on the 111 individuals of the ruminant Austeria / Marshallia elegans collected above. Standard amplification procedures were used: pre-denaturation at 94°C for 5min; denaturation at 94°C for 30S, annealing at 50°C for 30S, extension at 72°C for 1min, a total of 38 cycles; extension at 72°C for 7min, and storage at 4°C. After the amplified product was detected b...

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Abstract

The invention relates to the field of molecular biology, in particular to an ostertagia circumcincta / marshallagia circumcincta COI gene amplification primer and application thereof. The ostertagia circumcincta / marshallagia circumcincta COI gene amplification primer is shown as SEQ ID No.1 and SEQ ID No.2, and can be used for effectively amplifying out an ostertagia circumcincta / marshallagia circumcincta COI target fragment which cannot be amplified by universal primers LCO1490 and HCO2198 in the prior art. The species molecular identification requirements of ruminant ostertagia circumcincta / marshallagia circumcincta can be met; meanwhile, the reliable technical guidance can also be provided for the ostertagia circumcincta / marshallagia circumcincta phylogenetic relation study, and ruminant gastrointestinal parasitic disease prevention and control.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to an amplifying primer for the COI gene of Osteria / Marshallia elegans and its application. Background technique [0002] Ruminant gastrointestinal parasitic nematodes (gastrointestinal nematodes inruminants) include Trichostrongyloidae (Trichostrongyloidae) subfamily (Trichostrongylinae) and long thorn nematode subfamily (Mecistocirrinae) animal nematodes. Among them, Trichostrongylus subfamily Ostertagia, Marshallagia, Teladorsagia, Haemonchus, Cooperia, Trichostrongylus The 6 genera including Trichostrongylus are more harmful to the grazing livestock in northern pastoral areas of my country, especially Osteria and Marshall's nematodes. Osteria / Marshall's nematodes are endoparasites of the digestive tract, mainly in the fourth stomach (abomasum) and the folds of the intestinal tract of ruminant livestock. The above-mentioned parasites cause great losses to my country's animal hus...

Claims

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Application Information

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IPC IPC(8): C12N15/11C12Q1/68
Inventor 吕继洲吴绍强林祥梅袁向芬张永宁
Owner CHINESE ACAD OF INSPECTION & QUARANTINE
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