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Application of serum ssc-miR-215 as molecular marker on detecting stress injury of intestinal tract of pigling

A technology of ssc-mir-215 and intestine, which is applied in the field of detection of molecular markers of intestinal stress injury in piglets, and can solve problems such as lack of diagnostic value

Active Publication Date: 2015-10-14
ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the detection of the changes of the above indicators are all relative values, and the corresponding control samples are required as a reference and the conclusion can be drawn by analyzing the significant difference of the data, so it has no diagnostic value

Method used

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  • Application of serum ssc-miR-215 as molecular marker on detecting stress injury of intestinal tract of pigling
  • Application of serum ssc-miR-215 as molecular marker on detecting stress injury of intestinal tract of pigling
  • Application of serum ssc-miR-215 as molecular marker on detecting stress injury of intestinal tract of pigling

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Example 1: In this example, ssc-miR-215 was obtained by analyzing the correlation between serum microRNA and intestinal microRNA of weaned piglets, including the following steps:

[0023] 1. Extraction of total RNA from intestinal samples of piglets

[0024] 24 piglets (6 piglets per litter) were selected from 4 litters of 25-day-old piglets with the same parity and birth date as the research objects, and were divided into lactation group and weaning group (weaning on the same day) on average. 1d, 4d and 7d after weaning piglets in the weaning group, respectively, 4 piglets were slaughtered in each group (1 piglet per litter). The jejunal tissue samples were collected, and the total RNA samples of piglet intestinal tissues were extracted by Trizol method.

[0025] 2. Small RNA library construction

[0026] The total RNA samples of 4 biological repetitions of each treatment group were converted according to the concentration and mixed in equal amounts to form RNA sampl...

Embodiment 2

[0050] Example 2: In this example, the ssc-miR-215 obtained in Example 1 was used to mark intestinal stress injury in weaned piglets.

[0051] (1) Randomly select 24 weaned piglets and mark them as Nos. 1-24 respectively, collect whole blood of weaned piglets, and prepare serum samples No. 1-24;

[0052] (2) Total RNA in serum samples was extracted by Plasma / Serum Exosome Kit (Norgen, 49200) reagent;

[0053] (3) Using miRNeasy Mini Kit (Qiagen 217004) kit to extract serum microRNA in Total RNA;

[0054] (4) Use the ssc-miR-215 reverse transcription primer shown in SEQ ID NO.2 to reverse transcribe the microRNA to obtain cDNA: the reverse transcription system is: microRNA sample 2.5ul (40ng / ul), ssc-miR-215 reverse transcription primer 0.5ul (2uM), dNTP 0.25ul (10mM each), 5×RT buffer 1.0ul, RNase inhibitor 0.25ul (40U / ul), M-MLV 0.5ul (200U / ul) , the total volume of the reaction system is 5.0ul, place the uniformly mixed reaction system at 42°C for 60min, then place at 70°C...

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Abstract

The invention belongs to the field of biology, and relates to an application of serum ssc-miR-215 as a molecular marker on detecting stress injury of the intestinal tract of a pigling. A nucleotide sequence of serum ssc-miR-215 is represented as SEQ ID NO.1. A nucleotide sequence of a stem-loop primer required by reverse transcription of ssc-miR-215 is represented as SEQ ID NO.2. A nucleotide sequence of a forward primer required by augmentation of ssc-miR-215 is represented as SEQ ID NO.3. Diagnosis of stress injury of the intestinal tract of a weaned pigling is realized by adopting the above primers. A diagnostic method comprises extraction of serum total RNA, separation of small molecular RNA, synthesis of cDNA, and augmentation and authentication of PCR. The invention provides guidance for prevention, diagnosis and treatment of stress injury of the intestinal tract of a pigling, and provides a novel molecular marker for marker assisted selection of an anti-stress pig line.

Description

technical field [0001] The invention belongs to the technical field of livestock genetic engineering, and in particular relates to the application of serum ssc-miR-215 as a molecular marker for detecting intestinal stress injury in piglets. Background technique [0002] Piglet weaning stress is a major problem in pig farming worldwide, causing huge economic losses to the pig industry every year. Piglet diarrhea caused by stress is the most common and most harmful stress disease in pig production. The main target of weaning stress is the piglet's gastrointestinal tract, where damage to the small intestine is particularly severe. Weaning stress-induced tissue damage and remodeling of the piglet small intestine seriously affects the development and maturation of the piglet gut, thereby restricting piglet health and growth. Similarly, intestinal damage caused by weaning stress, resulting in a decline in the synthesis and absorption of gastrointestinal enzymes, is considered to...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/6883C12Q2600/166C12Q2600/178
Inventor 陶新徐子伟
Owner ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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