Duck beak atrophy and dwarfish syndrome nano PCR (polymerase chain reaction) detection kit and detection method thereof
A kit and syndrome technology, applied in the field of detection of duck short beak and long tongue syndrome, can solve problems such as insufficient amplification efficiency and non-specific product amplification in complex systems, so as to improve specific amplification yield and reduce dwell time. time, and the effect of improving detection efficiency
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0028] Embodiment 1 A kind of nano-PCR kit best embodiment that detects duck short beak and long tongue syndrome
[0029] The kit in this embodiment includes: (1) DNA extraction reagents for duck short beak and long tongue syndrome: proteinase K, lysate, sodium acetate, saturated phenol? chloroform? isoamyl alcohol, absolute ethanol, sterile distilled water.
[0030] (2) Nano-PCR reaction reagents: 2×NanoPCRMix, upstream primers, downstream primers; (4) Others: positive control and nuclease-free water. Among them, 2×NanoPCRMix is composed of DNA polymerase, 2×NanoPCR buffer and dNTP Mixture, and the positive control is the allantoic fluid of 9-11-day-old duck embryos after inoculation of Duck Short Beak Long Tongue Syndrome Virus, and the collection of 96-144 hours infected duck embryos , the primers are lyophilized powder, pure by HPLC. The primer sequences included in the kit of this example are shown in Table 1.
[0031] Table 1 Primer Sequence
[0032] Prime...
Embodiment 2
[0033] Embodiment 2 Non-diagnostic purpose detects the method for duck short beak and long tongue syndrome virus with nano-PCR method
[0034] The detection method of this example uses the kit in Example 1. The spleen and liver of diseased ducks were taken as samples to be tested.
[0035] The detection method of this embodiment comprises the following steps:
[0036] 1. The specific steps of virus DNA extraction are as follows: (1) After cutting the sample to be tested, add physiological saline according to the mass volume ratio of 1:5 and grind evenly, centrifuge at 3000-5000rpm for 5-10 minutes, take the supernatant clear. (2) Take 300 μL of the above-mentioned treated sample and positive control, add 10 μL of proteinase K, 300 μL of cell lysate, incubate at 50°C for 2 hours, add 610 μL of tris-balanced phenol, shake fully for 5 minutes, and centrifuge at 12,000 rpm for 10 minutes. (3) Transfer the supernatant to a new 1.5mL centrifuge tube, add an equal volume of phenol...
Embodiment 3
[0040]Example 3 Sensitivity Test of Nano-PCR Detection Method for Duck Short Beak and Long Tongue Syndrome
[0041] The nucleic acid template with a total DNA concentration of 225 ng / μL was calculated for duck short beak and long tongue syndrome virus by ultra-micro spectrophotometer measurement for sensitivity test. At the same time, nano-PCR and common PCR methods were used to detect the 10-fold diluted duck short beak and long tongue syndrome nucleic acid template.
[0042] The nano-PCR of duck short beak and long tongue syndrome virus and common PCR operation process are carried out according to embodiment 2, wherein, common PCR reaction changes 2 * NanoPCRMix in step 3 into 2 * PCR reaction solution, and its composition is as follows: DNA polymerase, 2×PCRbuffer, dNTPMixture, and other operating procedures remain unchanged.
[0043] The PCR amplification products were detected by 1% agarose gel electrophoresis, and the results were as follows: figure 2 shown. It can b...
PUM
| Property | Measurement | Unit |
|---|---|---|
| particle diameter | aaaaa | aaaaa |
Abstract
Description
Claims
Application Information
Login to View More