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Application of PSAP gene in preparation of product for detecting embryo death in early stage of pig pregnancy

A technology for early pregnancy and embryos, which is applied in the application field of PSAP gene in the preparation and detection of early pregnancy embryo death products in pigs, and can solve the problems of indistinguishability and errors.

Inactive Publication Date: 2021-05-14
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the observation method often causes some unavoidable errors: for example, the errors caused by the operator itself or the errors caused by the fact that some developing embryos in the first trimester have died or are about to die but are still difficult to distinguish with the naked eye at the time of slaughter.

Method used

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  • Application of PSAP gene in preparation of product for detecting embryo death in early stage of pig pregnancy
  • Application of PSAP gene in preparation of product for detecting embryo death in early stage of pig pregnancy
  • Application of PSAP gene in preparation of product for detecting embryo death in early stage of pig pregnancy

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1 Differential gene screening and immune response verification

[0036] 1. Experimental design and sample collection

[0037] A breeding farm selected 4 sows (Tibetan pigs) with similar body condition and normal development for artificial insemination. On the 28th day of pregnancy, the local slaughterhouse slaughtered the 4 sows, and quickly took out the uteri and put them in an ice box for delivery. to the lab. Subsequently, the uterus was opened longitudinally along the transmedial side of the uterus, and healthy and arrested embryos obtained from 4 different sows (eg, figure 1 shown), samples of healthy developing embryo site endometrium (HE) and arrested developing embryo site endometrium (AE) were collected, immediately immersed in liquid nitrogen and transferred to a -80°C freezer for subsequent RNA extraction.

[0038] 2. RNA extraction and Illumina sequencing of endometrial tissue

[0039] Total RNA was extracted from endometrial tissue using TRIzol r...

Embodiment 2

[0048] Example 2 PSAP gene selection and qRT-PCR verification

[0049] Since the spontaneous death of pig embryos during early pregnancy is related to the immune response at the maternal-fetal interface, a PSAP gene related to immune response was selected from 2357 differentially expressed genes as a candidate molecular marker for spontaneous death of pig embryos in early pregnancy , and by fluorescent quantitative PCR (qRT-PCR) to verify the difference in the expression of PSAP in embryonic normal site (HE) and developmental arrest site (AE).

[0050] (1) cDNA synthesis

[0051] The extracted qualified total RNA of HE and AE samples was used to synthesize cDNA, and the reverse transcription was performed using the Takara reverse transcription kit (PrimeScript TM RT reagent Kit with gDNA Eraser (Perfect RealTime)), first remove genomic DNA, the total reaction system is 10 μl (Table 1), the reaction program is: 42 ° C for 2 min, 4 ° C to stop the reaction; then cDNA synthesis...

Embodiment 3

[0068] Example 3 Independent sample qRT-PCR verification of early pregnancy pig embryo spontaneous death molecular marker PSAP

[0069] In a group of sows with independent samples, get 2 gestational 35-day sows (Tibetan pigs) healthy development embryo site endometrium (HE) and embryo death degeneration site endometrium (AE) samples, refer to the embodiment 2. Extract the total RNA of the sample and perform qRT-PCR analysis. For the specific reaction system and procedure, refer to the quantitative part of qRT-PCR in Example 2.

[0070] The result is as Figure 5 As shown, there was also a significant difference in PSAP gene expression between HE and AE samples at 35 days of pregnancy. The results showed that the PSAP gene can be used as a molecular marker for the spontaneous death of pig embryos in early pregnancy.

[0071] For the sample to be tested, with reference to Example 2, the total RNA of the sample was extracted and analyzed by qRT-PCR, and the endometrium (HE) sam...

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Abstract

The invention relates to the technical field of biomolecular markers, in particular to application of a PSAP gene in preparation of a product for detecting embryo death in an early stage of pig pregnancy. By means of Illumina sequencing, differential gene screening, bioinformatics analysis, qRT-PCR and other methods, it is determined that spontaneous death of porcine embryogenesis in the early pregnancy process is related to immunoreaction activity weakening of a maternal-fetal interface, and it is further screened and determined that the PSAP gene can serve as the early pregnancy porcine embryo spontaneous death molecular marker. Compared with a traditional observation method, the PSAP gene can be used for more accurately measuring spontaneously dead embryos in the early pregnancy process, and a new means is provided for scientific researchers for researching regulation and control of the embryonic development process and molecular mechanisms such as regulation and control of embryonic spontaneously dead and endometrium regulation and control.

Description

technical field [0001] The invention relates to the technical field of biomolecular markers, in particular to the application of a PSAP gene in the preparation and detection of embryonic death products in the early pregnancy of pigs. Background technique [0002] The litter size of a sow determines the production efficiency of a sow to a certain extent. Therefore, the spontaneous death of embryos during sow pregnancy has always been concerned in the pig industry. How to reduce the spontaneous death of embryos and improve the quality of sows The litter size of piglets has become an urgent problem for scientific researchers in various countries. About 20-45% of pig embryos will die spontaneously at different stages of pregnancy, especially in the early pregnancy process, about 12 to 30 days of pregnancy, and 20-30% of pig embryos will die spontaneously. How to accurately distinguish whether the early embryonic development is normal is very important for researchers who study ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6883C12N15/11G01N33/68
CPCC12Q1/6883G01N33/689C12Q2600/158G01N2800/36
Inventor 吴珍芳洪林君臧旭鹏周臣王雯竞蔡更元顾婷杨杰杨化强郑恩琴
Owner SOUTH CHINA AGRI UNIV
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