Molecular marker for identifying duck feed utilization rate character based on ITGB1 gene as well as identification method and application thereof
By using a molecular marker identification method based on the ITGB1 gene, the problem of early identification of feed utilization traits in duck breeding has been solved, realizing a simple, rapid, and low-cost breeding method that improves feed efficiency in meat ducks.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2026-02-14
- Publication Date
- 2026-04-10
AI Technical Summary
In the poultry sector, especially in ducks, insufficient research on the ITGB1 gene has led to slow progress in conventional phenotypic breeding and selection, making it difficult to identify feed utilization traits in the early stages and affecting the improvement of feed efficiency in meat ducks.
Based on the ITGB1 gene, molecular markers were developed, and specific amplification primers were designed for PCR amplification and restriction endonuclease digestion. Combined with agarose gel electrophoresis detection, the feed utilization trait of ducks was identified, providing a simple, rapid, and low-cost molecular marker-assisted breeding method.
This method enables early selection of duck feed utilization traits, provides a new breeding approach, improves breeding efficiency, reduces costs, and is suitable for molecular marker-assisted breeding experiments.
Smart Images
Figure CN121826178A_ABST
Abstract
Description
Technical Field
[0001] This invention belongs to the field of molecular marker technology, specifically relating to a molecular marker for identifying duck feed utilization traits based on the ITGB1 gene, its identification method, and its application. Background Technology
[0002] Residual Feed Intake (RFI) is defined as the difference between an animal's actual feed intake and its expected feed intake based on its body size and growth. It is independent of production levels; a lower RFI indicates higher animal efficiency. Although not directly applied to breeding programs, it can guide the selection of valuable individuals within a population due to its lack of correlation with body weight and weight gain. While the Feed Conversion Ratio (FCR) is a non-normally distributed ratioistic trait with no true mean and variance, making it difficult to improve without direct statistical impact, RFI is not a ratioistic trait. Over decades of development, RFI has been widely applied to the genetic improvement of livestock feed efficiency because it has the advantage of reducing FI and improving feed efficiency without sacrificing body weight or body weight gain (BWG) (Ferreira J, Chay-Canul AJ, De Barbieri I, et al. Compilations and updates on residual feed intake in sheep[J]. Trop Anim Health Prod. 2024, 56(5):172.;ZouX, Liu T, Li Y, et al. Long-term divergent selection for residual feed intake in Chinese broiler chickens. Poult Sci. 2023, 102(3):102298.). Therefore, RFI can be used as a measure of feed utilization during breeding.
[0003] ITGB1 (integrin subunit β1) is a protein-coding gene and a core member of the integrin family. Its bidirectional signal transduction mechanism is key to cell adhesion and signal transduction (Hynes R O. Integrins: bidirectional, allosteric signaling machines[J]. cell, 2002, 110(6):673-687.). Transcriptome and metabolome studies in beef cattle have revealed that the ITGB1 gene is a key gene in fatty acid metabolism, fatty acid degradation, fatty acid elongation, and the PPAR signaling pathway. Studies on local pigs have found that the ITGB1 gene is a key gene affecting the body size difference between local pigs and hog pigs. On chromosome 10 of local pigs, the ITGB1 region shows a strong positive selection signal, and its expression level is significantly higher in the adipose tissue of local pigs than in hog pigs, and increases with age. This suggests that the ITGB1 gene may affect body size by regulating the lipid metabolism efficiency of adipose tissue (Yu H, Yu S, Guo J, et al. Comprehensive Analysis of Transcriptome and Metabolome Reveals Regulatory Mechanism of Intramuscular Fat Content in Beef Cattle). [J]. J Agric Food Chem. 2024, 72(6):2911-2924. WuG, Yu M, Liu T, et al. Integration of multiomics data reveals selection characteristics of ITGB1 that are associated with size differentiation inpigs[J]. International Journal of Molecular Sciences, 2025, 26(4):1569.) Studies have shown that the expression of the ITGB1 gene in the liver of Landes geese is significantly upregulated after force-feeding; further cell experiments have found that factors related to fatty liver formation, such as glucose, oleic acid, linoleic acid and palmitic acid, can induce the expression of ITGB1 in primary hepatocytes of geese; this suggests that the upregulation of the ITGB1 gene may be closely related to the formation of fatty liver (Gu W, Zhang Y, Zhang J, et al.).The relationship between ITGB1 gene expression and theformation of goose fatty liver[J]. 2021, 771-776). .
[0004] Current research on the ITGB1 gene mainly focuses on mammals and fish, with most studies revolving around disease research, followed by research on fatty acid metabolic regulation and lipid deposition mechanisms. However, in the poultry sector, especially in ducks, research on the ITGB1 gene is still insufficient. Therefore, it is crucial to explore the specific impact of ITGB1 gene variations and expression patterns on feed conversion ratio in broiler ducks and to reveal the underlying molecular mechanisms. This will not only help us to more comprehensively understand the key role of the ITGB1 gene in the growth, development, and feed utilization of broiler ducks, but also provide a strong theoretical basis for improving feed efficiency in broiler ducks, thus contributing to cost reduction and efficiency improvement and promoting the sustainable development of the poultry farming industry. Therefore, this invention proposes a molecular marker for identifying duck feed utilization traits based on the ITGB1 gene, along with its identification method and application. Summary of the Invention
[0005] The purpose of this invention is to provide a molecular marker for identifying duck feed utilization traits based on the ITGB1 gene, along with its identification method and application, in order to solve the above-mentioned problems. This invention targets SNP (single nucleotide polymorphism) molecular markers of candidate genes related to duck feed utilization traits, thereby addressing the challenge of slow progress in conventional phenotypic breeding and achieving early identification of feed utilization traits.
[0006] The present invention achieves the above objectives through the following technical solutions: A molecular marker for identifying duck feed utilization traits based on the ITGB1 gene, wherein the nucleotide sequence of the molecular marker is shown in SEQ ID NO.1, and the 600th base of the nucleotide sequence is C or T.
[0007] Application of a molecular marker based on the ITGB1 gene for identifying duck feed utilization traits.
[0008] As a further optimization of the present invention, if the molecular marker type of the duck to be tested is CT, the duck has the best feed utilization trait; if the molecular marker type of the duck to be tested is TT, the duck has a moderate feed utilization trait; and if the molecular marker type of the duck to be tested is CC, the duck has a poor feed utilization trait.
[0009] A method for identifying duck feed utilization traits using molecular markers as described above includes the following steps: (1) Extract total DNA from the venous blood of the ducks to be tested; (2) Design specific amplification primers with the molecular marker site and the sequence composed of its upstream and downstream bases as the target sequence, use the total DNA as a template, and perform PCR amplification using the specific amplification primers to obtain the amplification product; (3) Genotyping and sequencing of the amplification products to obtain the molecular marker type of the duck to be tested; (4) Determine the duck feed utilization rate trait based on the molecular marker type.
[0010] As a further optimization of the present invention, the sequence of the specific amplification primers is as follows: SEQ ID NO.2: Forward primer: CAGAAATGATACCCACGCC; SEQ ID NO.3: Reverse primer: CAAACACCGAACAACCAAAG.
[0011] As a further optimization of the present invention, the genotyping detection method involves obtaining enzyme digestion products by digesting the amplification products with restriction endonucleases, detecting the enzyme digestion products using agarose gel electrophoresis, and performing genotyping based on the images. If the enzyme digestion products: If it contains one stripe, it is of type TT; If it contains two stripes, it is of type CC; If it contains 3 bands, it is a CT type.
[0012] As a further optimization of the present invention, the amplification product is digested with HpyCH4IV restriction endonuclease.
[0013] As a further optimization of the present invention, if the molecular marker type of the duck to be tested is CT, the duck has the best feed utilization trait; if the molecular marker type of the duck to be tested is TT, the duck has a moderate feed utilization trait; and if the molecular marker type of the duck to be tested is CC, the duck has a poor feed utilization trait.
[0014] As a further optimization of the present invention, the enzyme digestion products are detected by agarose gel electrophoresis at a concentration of 1.5%-2.0%.
[0015] The present invention has the following beneficial effects: This invention develops a molecular marker based on research into the relationship between the ITGB1 gene and duck feed utilization traits. By identifying the types of these molecular markers present in the duck genome, selection of ducks for feed utilization traits can be achieved, establishing a breeding method for early selection of poultry feed utilization. This provides a new molecular marker-assisted breeding method for detecting duck growth traits. This method is simple, rapid, low-cost, and does not require special instruments, making it suitable for the needs of molecular marker-assisted breeding experiments. Attached Figure Description
[0016] Figure 1 Agarose gel electrophoresis images of PCR amplification products from a portion of the samples; Figure 2 Agarose gel electrophoresis image of the enzyme digestion products obtained by enzyme digestion of PCR amplification products from a portion of the samples; Figure 3 This is the genotype verification sequencing result for the C144606873T site (the 600th base in SEQ ID NO.1) in the duck ITGB1 gene. Detailed Implementation
[0017] The present application will now be described in further detail with reference to the accompanying drawings. It should be noted that the following specific embodiments are only used to further illustrate the present application and should not be construed as limiting the scope of protection of the present application. Those skilled in the art can make some non-essential improvements and adjustments to the present application based on the above application content.
[0018] 1. Materials Unless otherwise specified, the methods used in this embodiment are conventional methods known to those skilled in the art, and the reagents and materials used are commercially available products.
[0019] 2. Method 2.1 Primer Design The DNA sequence corresponding to the ITGB1 gene (Gene ID: NC_092588.1) shown in SEQ ID NO.1 was found in the duck genome database. Using the partial sequence of the ITGB1 gene DNA (the sequence consisting of the molecular marker site and its upstream and downstream bases) as a template, specific amplification primers were designed. The specific amplification primer sequences are shown below: SEQ ID NO.2: Forward primer: CAGAAATGATACCCACGCC; SEQ ID NO.3: Reverse primer: CAAACACCGAACAACCAAAG.
[0020] The amplifiable region of the primer is 845 bp in length, and the amplified sequence is shown in SEQ ID NO.4, which contains a molecular marker of the C / T mutation at the C144606873T site (the 600th base in SEQ ID NO.1).
[0021] 2.2 Extraction of total DNA from blood A total of 549 Qiangying ducks were selected, and blood was collected from the medial metatarsal vein. Total DNA was extracted from the blood. The total DNA was extracted from the blood samples from the medial metatarsal vein using a blood DNA extraction kit produced by Tiangen Biotech Co., Ltd. The extraction steps were performed according to the kit instructions.
[0022] 2.3 PCR Amplification Using Mix produced by Shanghai Yisheng Biotechnology Co., Ltd., PCR amplification of the target fragment of the ITGB1 gene was performed using pre-synthesized sequencing-specific primers. The PCR amplification system is shown in Table 1. Table 1 PCR amplification system The PCR reaction conditions were as follows: 95℃ pre-denaturation for 5 min; first step: 95℃ denaturation for 45 s; second step: 60℃ annealing for 45 s (annealing temperature is set according to the primers); third step: 72℃ extension for 30 s, with the second and third steps repeated 34 times for a total of 35 cycles; and a final extension at 72℃ for 10 min.
[0023] 2.4 Detection and Sequencing of PCR Amplification Products PCR amplification products were detected using 2% agarose gel electrophoresis, such as... Figure 1 As shown, after imaging with a gel imaging system, a band of approximately 845 bp in length was obtained, which is consistent with the predicted length, indicating that the target fragment was obtained. The PCR product was sent to Beijing Qingke Biotechnology Co., Ltd. (Nanjing), and the sequence is shown in SEQ ID NO.4, which is consistent with the predicted result.
[0024] 2.5 Genotyping 2.5.1 Prepare the enzyme digestion system as shown in Table 2. The enzyme digestion conditions are 37℃, PCR overnight, and digest the PCR amplification product using HpyCH4IV restriction endonuclease [restriction site is A^CGT] from Hefei Ruijie Technology Co., Ltd. Table 2 Enzyme digestion system 2.5.2. Detection was performed using 1.5% mass ratio low-voltage agarose gel electrophoresis to obtain results such as... Figure 2 The results shown are partial; among them, if the enzyme digestion product contains 1 band, it is of the TT type; contains 2 bands, it is of the CC type; and contains 3 bands, it is of the CT type.
[0025] 2.6. Enzyme digestion and sequencing verification Statistical analysis of the gene genotyping agarose gel electrophoresis images yielded three genotypes: TT, CC, and CT. One individual was selected from each of these three genotypes for sequencing alignment. The sequencing alignment images are shown below. Figure 3 As shown in the sequencing results, C mutated to T, and the arrows indicate the mutation locations, which is consistent with the enzyme digestion typing results.
[0026] 2.7 Effect Verification To determine the association between the C / T polymorphism at the C144606873T locus of the duck ITGB1 gene and important phenotypic traits in ducks, 549 Qiangying ducks from step 2.2 above were used as experimental materials. Feed intake (ADFI), average daily gain (ADG), and metabolite weight gain (MBW) were recorded from 21 to 38 days of age. 0.75 The feed conversion ratio (FCR) and residual feed intake (RFI) were used to genotype 549 Qiangying ducks using the genotyping method described in step 2.5 above. The results are shown in Table 3. Table 3. Genotype detection results for individuals with different phenotypes Experimental conclusion: The chi-square test results showed that the genotypes of the experimental duck population were in Hardy-Weinberg equilibrium (P>0.05).
[0027] 2.8 Statistical Analysis The association between the three genotypes and the duck feed utilization trait was analyzed using SPSS software analysis of variance. The association analysis results between different genotypes and each trait are shown in Table 4. Table 4. Association analysis between duck ITGB1 genotype and duck feed utilization trait. Note: Different lowercase letters in the same row indicate significant differences (P<0.05), and different uppercase letters in the same row indicate extremely significant differences (P<0.01).
[0028] Experimental Conclusions: As shown in Table 4, for the ITGB1 gene C144606873T locus, the daily feed intake (ADFI) of CT genotype individuals was significantly lower than that of CC genotype individuals (P<0.05); the feed conversion ratio (FCR) and residual feed intake (RFI) of CT genotype individuals were extremely significantly lower than those of CC genotype individuals (P<0.01), and the RFI of TT genotype individuals was also extremely significantly lower than that of CC genotype individuals (P<0.01); the three genotypes showed similarities in average daily gain (ADG) and metabolite weight gain (MBW). 0.75There were no significant differences in feed utilization in the CT genotype, which leads to the conclusion that individuals with the CT genotype had the best feed utilization trait, individuals with the TT genotype had a moderate feed utilization trait, and individuals with the CC genotype had a poor feed utilization trait.
[0029] The above embodiments merely illustrate several implementation methods of the present invention, and their descriptions are relatively specific and detailed, but they should not be construed as limiting the scope of the present invention. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these all fall within the protection scope of the present invention.
Claims
1. A molecular marker for identifying duck feed utilization traits based on the ITGB1 gene, characterized in that, The nucleotide sequence of the molecular marker is shown in SEQ ID NO.1, wherein the 600th base of the nucleotide sequence is C or T.
2. The application of the molecular marker as described in claim 1 in identifying duck feed utilization traits.
3. The application according to claim 2, characterized in that, If the molecular marker type of the duck being tested is CT, the duck has the best feed utilization trait; if the molecular marker type of the duck being tested is TT, the duck has a moderate feed utilization trait; if the molecular marker type of the duck being tested is CC, the duck has a poor feed utilization trait.
4. A method for identifying duck feed utilization traits using the molecular markers as described in claim 1, characterized in that, Includes the following steps: (1) Extract total DNA from the venous blood of the ducks to be tested; (2) Design specific amplification primers with the molecular marker site and the sequence composed of its upstream and downstream bases as the target sequence, use the total DNA as a template, and perform PCR amplification using the specific amplification primers to obtain the amplification product; (3) Genotyping and sequencing of the amplification products to obtain the molecular marker type of the duck to be tested; (4) Determine the duck feed utilization rate trait based on the molecular marker type.
5. The method for identifying duck feed utilization traits using molecular markers according to claim 4, characterized in that, The sequence of the specific amplification primers is as follows: SEQ ID NO.2: Forward primer: CAGAAATGATACCCACGCC; SEQ ID NO.3: Reverse primer: CAAACACCGAACAACCAAAG.
6. The method for identifying duck feed utilization traits using molecular markers according to claim 4, characterized in that, The genotyping detection method involves obtaining restriction endonuclease digestion of the amplification products, detecting the digested products using agarose gel electrophoresis, and performing genotyping based on the images. If the digested products: If it contains one stripe, it is of type TT; If it contains two stripes, it is of type CC; If it contains 3 bands, it is a CT type.
7. The method for identifying duck feed utilization traits using molecular markers according to claim 6, characterized in that, The amplified product was digested with HpyCH4IV restriction endonuclease.
8. The method for identifying duck feed utilization traits using molecular markers according to claim 6, characterized in that, If the molecular marker type of the duck to be tested is CT, the duck has the best feed utilization trait; if the molecular marker type of the duck to be tested is TT, the duck has a moderate feed utilization trait; if the molecular marker type of the duck to be tested is CC, the duck has a poor feed utilization trait.
9. The method for identifying duck feed utilization traits using molecular markers according to claim 6, characterized in that, The enzyme digestion products were detected by agarose gel electrophoresis at a concentration of 1.5%–2.0%.