Application of polypeptide in feed additive for reducing transport stress reaction of pigs

Abstract

The invention provides application of polypeptide in a feed additive for reducing pig transport stress response, and belongs to the technical field of feed. In-vitro experiments find that the porcine granulation-promoting regulatory factor WOB19092.1 polypeptide has an obvious promoting effect on the cell viability of porcine small intestine epithelial cells, and can improve the oxidative stress of the porcine small intestine epithelial cells caused by ultraviolet lamp irradiation, reduce the MDA level of the cells and improve the SOD level of the cells. Clinical experiments show that the pig granulation-promoting regulatory factor WOB19092.1 polypeptide serving as the feed additive for reducing the transport stress response of the pigs effectively reduces the death rate and morbidity caused by transport stress of the piglets, obviously improves the anorexia condition of the piglets caused by the transport stress, and promotes weight gain of the piglets in the initial stage of entering a fattening pig farm.

Description

Technical Field

[0001] The present invention belongs to the technical field of feeds, and particularly relates to the application of a polypeptide in a feed additive for reducing the transportation stress response of pigs. Background Art

[0002] With the development of breeding technology, the pig breeding mode in China has developed from traditional rural household scattered breeding to large-scale intensive breeding. The number of pigs sold in a batch by large-scale pig farms often reaches several thousand to tens of thousands. Most pig farms mainly focus on professionally raising fattening pigs, purchasing piglets from breeding farms and further fattening them until they are sold. The purchased piglets are usually transported to fattening pig farms by trucks, and the distance between breeding farms and fattening pig farms is often far, sometimes requiring long-distance transportation. Piglets are loaded and sent from breeding farms to fattening pig farms a few days after weaning. Due to the small age and poor stress resistance of piglets themselves, long-distance transportation will cause serious stress responses, resulting in illness and death during transportation, and problems such as anorexia and growth stagnation for several days after being transported to fattening pig farms. The economic losses caused by the high incidence, high mortality and long-term growth stagnation of piglets due to transportation stress responses are the top problems for fattening pig farms. Therefore, how to reduce the transportation stress response of piglets by means of drugs or feed additives is an urgent problem to be solved currently.

[0003] Bioactive peptides are small molecule active peptides directly formed by organisms or produced by the degradation of protein molecules. Bioactive peptides are usually closely related to processes such as nutrition, hormones, immune regulation, antibacterial, antiviral, and antioxidant. Summary of the Invention

[0004] The purpose of the present invention is to provide the application of a polypeptide in a feed additive for reducing the transportation stress response of pigs, belonging to the technical field of feeds.

[0005] To achieve the above purpose, the present invention provides the following technical solutions: First, the present invention provides the application of a polypeptide in a feed additive for reducing the transportation stress response of pigs, and the polypeptide is porcine myogenic regulatory factor WOB19092.1 polypeptide.

[0006] Further, the amino acid sequence of the porcine myogenic regulatory factor WOB19092.1 polypeptide is RGLVMTAKEGGTNIDS.

[0007] Further, the feed additive can be used to reduce the incidence and mortality caused by transportation stress in piglets.

[0008] Further, the feed additive can be used to improve the anorexia situation of piglets caused by transportation stress and promote the weight gain of piglets in the initial stage after entering fattening pig farms.

[0009] Second, the present invention provides a polypeptide for promoting the cell viability of porcine small intestinal epithelial cells. The polypeptide is porcine myogenic regulatory factor WOB19092.1 polypeptide, and the amino acid sequence of the porcine myogenic regulatory factor WOB19092.1 polypeptide is RGLVMTAKEGGTNIDS.

[0010] Third, the present invention provides a polypeptide for reducing the oxidative stress response of porcine small intestinal epithelial cells. The polypeptide is porcine myogenic regulatory factor WOB19092.1 polypeptide, and the amino acid sequence of the porcine myogenic regulatory factor WOB19092.1 polypeptide is RGLVMTAKEGGTNIDS.

[0011] Beneficial effects: Through in vitro experiments, the present invention found that porcine myogenic regulatory factor WOB19092.1 polypeptide has an obvious promoting effect on the cell viability of porcine small intestinal epithelial cells, and can improve the oxidative stress of porcine small intestinal epithelial cells caused by ultraviolet lamp irradiation, reduce the cell MDA level, and increase the cell SOD level. Therefore, it can be used to prepare a feed additive for reducing porcine transportation stress response.

[0012] Secondly, through clinical experiments, the present invention found that porcine myogenic regulatory factor WOB19092.1 polypeptide, as a feed additive for reducing porcine transportation stress response, effectively reduces the mortality rate and morbidity rate caused by porcine transportation stress, and significantly improves the anorexia situation of piglets caused by transportation stress, and promotes the weight gain of piglets in the initial stage of entering the fattening pig farm. The weight gain of piglets in the first month increased by 33.45% compared with the control group. Feeding porcine myogenic regulatory factor WOB19092.1 polypeptide feed additive one day before piglet transportation and in the first three days before transporting from the breeding pig farm to the fattening pig farm can effectively improve the breeding efficiency of the fattening pig farm. Description of the Drawings

[0013] Figure 1 Mass spectrometry detection results of porcine myogenic regulatory factor WOB19092.1 active polypeptide; Figure 2 Influence of the polypeptide on the cell viability of porcine small intestinal epithelial cells; Figure 3 Influence of the polypeptide on the MDA level of porcine small intestinal epithelial cells; Figure 4 Influence of the polypeptide on the SOD level of porcine small intestinal epithelial cells. Detailed Embodiments

[0014] Example 1: Mining of the active polypeptide sequence of porcine myogenic regulatory factor WOB19092.1 Analyze the amino acid sequence of porcine myogenic regulatory factor WOB19092.1, truncate it from the α-helix region, select a 15-amino acid peptide segment at positions 15 - 30, and its amino acid sequence is: RGLVMTAKEGGTNIDS. Entrust Sangon Biotech (Shanghai) Co., Ltd. to synthesize it. Figure 1 It is the mass spectrometry detection result of the synthesized polypeptide.

[0015] Example 2: Detect the effect of the polypeptide on the cell viability of porcine intestinal epithelial cells (1) Inoculate porcine intestinal epithelial cells in the logarithmic growth phase into a 96-well plate at a density of 1×10 4 cells per well; (2) When the cells are cultured in a cell incubator until the cell density is close to 80%, add DMEM medium containing the polypeptide at final concentrations of 0 μg / ml, 20 μg / ml, 40 μg / ml, 80 μg / ml, 160 μg / ml, 320 μg / ml, and 640 μg / ml. Set 4 parallels for each group, and set a blank control group at the same time; (3) After culturing in a cell incubator for 24 h, use CCK-8 to measure the OD450 value and calculate the cell survival rate. The cell survival rate = (OD450 of the experimental group - OD450 of the blank control group) / (OD450 of the control group - OD450 of the blank control group).

[0016] It can be seen from Figure 2 that within the concentration range of 20 μg / ml - 640 μg / ml, the synthesized polypeptide has an obvious promoting effect on the cell viability of porcine intestinal epithelial cells. The promoting effect from 20 μg / ml to 160 μg / ml increases with the increase of the polypeptide concentration, and it no longer increases from 160 μg / ml to 640 μg / ml.

[0017] Example 3: Detect the effect of the polypeptide on the oxidative stress of porcine intestinal epithelial cells (1) Inoculate porcine intestinal epithelial cells in the logarithmic growth phase into a 96-well plate at a density of 2×10 5 cells per well; (2) When the cells are cultured in a cell incubator until the cell density is close to 80%, add DMEM medium containing the synthesized polypeptide at final concentrations of 0 μg / ml, 20 μg / ml, 40 μg / ml, 80 μg / ml, 160 μg / ml, 320 μg / ml, and 640 μg / ml. Set 4 parallels for each group, and set a blank control group at the same time; (3) The control group is continued to be cultured in a cell incubator for 24 h; the cells in the oxidative stress group are placed in a biosafety cabinet and irradiated with ultraviolet light for 30 minutes, and then transferred to a cell incubator and continued to be cultured for 24 h; (4) Remove the culture medium, wash the cells with PBS, and then use superoxide dismutase (SOD) and malondialdehyde (MDA) assay kits to detect the contents of SOD and MDA.

[0018] From Figure 3 It can be seen that within the concentration range of 20 μg / ml - 640 μg / ml, the synthetic polypeptide can significantly reduce the MDA level in porcine intestinal epithelial cells. The effect reaches the maximum at 320 μg / ml, and the effect of 640 μg / ml is equivalent to that of 320 μg / ml. From Figure 4 It can be seen that within the concentration range of 20 μg / ml - 640 μg / ml, the synthetic polypeptide can significantly increase the SOD level in porcine intestinal epithelial cells. The effect reaches the peak at 320 μg / ml, and the effect of 640 μg / ml is equivalent to that of 320 μg / ml. Ozone generated by ultraviolet lamp irradiation causes oxidative stress in porcine intestinal epithelial cells. After adding the polypeptide to the culture medium, the MDA level is significantly reduced and the SOD level is significantly increased, and the oxidative stress response is improved. Therefore, treating porcine intestinal epithelial cells with this polypeptide can effectively reduce the oxidative stress caused by ultraviolet lamp irradiation.

[0019] Example 4: Effect of porcine myogenic regulatory factor WOB19092.1 polypeptide on transportation stress in piglets (1) Add porcine myogenic regulatory factor WOB19092.1 polypeptide No. 3 to the basal diet of piglets in different proportions and stir well. Prepare three kinds of feeds according to the weight ratio of polypeptide feed additive: basal diet, 1:1000, 1:10000, 1:100000; (2) Select 400 piglets at 21 days of age, with a body weight of 7 ± 0.2 kg, weaned at 21 days of age, randomly divided into 4 groups, 100 pigs per group, and raised in isolation; (3) Feed once at 8:00 and 16:00 on the day of weaning, and allow free access to food and water. The experimental groups were fed with feeds containing polypeptide feed additives, and the control group was only fed with basal diet. After one day of feeding, the pigs were transported 200 km by car to the fattening pig farm at 22 days of age, and the experimental groups continued to be fed with feeds containing polypeptide feed additives for 3 days. After 3 days, the polypeptide feed additive was removed, and all experimental pigs were fed with basal diet for 27 days. During this period, the food intake and mental state of the experimental pigs were observed daily, and the survival rate, morbidity and weight gain of pigs in each group were counted at the end of the experiment.

[0020] Table 1 Effect of feeding with polypeptide feed additives at different proportions on the growth of piglets

[0021] As can be seen from Table 1, when the porcine myogenic regulatory factor WOB19092.1 polypeptide of the present invention is added to the basal diet at a ratio of 1:1000 and 1:10000 as a feed additive to feed piglets, it can improve the survival rate, reduce the incidence of diseases, and reduce the direct losses caused by long-distance transportation stress to piglets. Moreover, it significantly improves the anorexia of piglets caused by transportation stress, promotes the weight gain of piglets in the initial stage of entering the fattening pig farm, and the weight gain of piglets in the first month is increased by 33.45% compared with the control group.

Claims

1. Use of a polypeptide in a feed additive for reducing pig transportation stress response, characterized in that: The polypeptide is a porcine myogenic regulatory factor WOB19092.1 polypeptide, and the amino acid sequence of the porcine myogenic regulatory factor WOB19092.1 polypeptide is RGLVMTAKEGGTNIDS.

2. The use according to claim 1, characterized in that: The feed additive can be used to reduce the morbidity and mortality caused by piglet transportation stress.

3. The use according to claim 1, characterized in that: The feed additive can be used to improve the anorexia of piglets caused by transportation stress and promote the initial weight gain of piglets in a fattening farm.

4. A polypeptide for promoting cell viability of porcine small intestinal epithelial cells, characterized in that: The polypeptide is a porcine myogenic regulatory factor WOB19092.1 polypeptide, and the amino acid sequence of the porcine myogenic regulatory factor WOB19092.1 polypeptide is RGLVMTAKEGGTNIDS.

5. A polypeptide for reducing oxidative stress response in porcine small intestinal epithelial cells, characterized in that: The polypeptide is a porcine myogenic regulatory factor WOB19092.1 polypeptide, and the amino acid sequence of the porcine myogenic regulatory factor WOB19092.1 polypeptide is RGLVMTAKEGGTNIDS.

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