Abstract: In this experiment, 100 100-day-old fattening piglets were selected as test subjects. The genetic polymorphism of myogenin (MyoG) gene was analyzed by PCR-SSCP technique, and the relationship between genotype and poached pork quality was studied. Sex. The results showed that there was a polymorphism at the exon 1-1 site of the MyoG gene. The polymorphism of MyoG gene has a great influence on the meat quality index of poached pigs and Landrace pigs, and reached a significant level.
Key words: pouch pig; MyoG gene; PCR-SSCP analysis; meat quality
Poached pigs are a kind of northeastern pigs, mainly distributed in Liaoning, Jilin and Heilongjiang provinces. The whole body is black, the winter is fluffy, the mane is of good quality and the body is strong. The face is long and straight, and the ear is drooping. The back is flat or slightly concave, the chest is deep, the limbs are thick and strong, and the body is lying. The hindquarters are narrow and inclined. Strong cold resistance, strong adaptability, resistance to rough feeding, good running, low incidence and good meat quality.
Myogenin (MyoG) is an important factor regulating skeletal muscle development, and MyoG is a member of the Myogenin determination gene (MyoD) gene family. The MyoD (also known as muscle regulatory factory, MRF) gene family encodes four distinct muscle-specific transcription factors: MyoD, MyoG, Myf5, and MRF4, which represent many aspects of controlling skeletal muscle production. Key adjustment factor. Among them, the MyoG gene is the only gene in the MyoD gene family that can be expressed in all skeletal muscle cell lines, and plays a key regulatory role. At present, the polymorphism of the MyoG gene in pigs has been studied abroad. For example, Ernst et al. (1998) first analyzed the genetic variation of the MyoG gene in pigs. They found that the 4.9 kb and 4.2 kb DNA fragments were polymorphic fragments. The separation of MyoG alleles containing 4.9 kb and 4.2 kb fragments in four second-generation and three-generation families was consistent with Mendel's separation law. The frequency of the MyoG allele of 83 pigs unrelated in 8 varieties was detected, and the frequency of MyoG alleles containing 4.9 kb and 4.2 kb DNA fragments was found to be 0.42% and 0.58%, respectively. TePas et al. (1996) found that the MyoG gene has three variant sites: one unique to Meishan pigs, one unique to Dutch Landrace and Duroc pigs, and the other is a variant site shared by all studied pigs; preliminary study These mutation sites were shown to be related to the number of pigs and the number of muscle fibers. In recent years, with the deepening of the research on the pig MyoG gene, some new progress has been made. As an important candidate for improving the quality of pork, the MyoG gene needs to be studied in depth.
1 Materials and methods
1.1 Materials
The high-quality pure-bred poached pigs and Changbai pigs used in this experiment were provided by Minsheng Hobao Pig Farm in Jianchang County, Huludao City. Blood was collected from the ear vein. All test pigs collected about 2 mL of blood. The blood was placed in a centrifuge tube with 10% EDTA anticoagulation and numbered separately. All of the test pigs were tested for MyoG genotype by PCR-SSCP analysis, and then selected for each genotype of pure-bred poached pigs weighing approximately 75 kg.
1.2 Primer design
The primers designed in this trial (Ann Soumillion, et al., 1996) were synthesized by Shenyang Baotek Bioengineering Co., Ltd. The primer sequence is:
Primer 1, R1: 5'-GGA ATT CCA TGG AGC TGT ATG AAC ATC-3'
F2: 5'-TCC GGC TCC TCC CTG GTT GA-3'
Primer 2, R1: 5'-TCA GGA AGA ACT GAA GGC TG-3'
F2: 5'-GTT TCC TGG GGT GTT GC -3'
1.3 PCR-SSCP analysis
PCR amplification system 50 μL. Amplification conditions: pre-denaturation temperature was 93 ° C, 3 min. Denatured at 93 ° C, 35 S, annealing temperature was 61 ° C, 30 S. Extend 30S at 72 °C.
Take the PCR product, add denaturing agent, paraffin oil, boil for 5min, take it out and put it into the ice bath for more than 8min, then all the samples, 10~15°C (specific temperature depends on the test, not too high). Start electrophoresis at 220V for 10min, then at 140V (voltage can not be higher than 120, otherwise it is possible to make the strip run) 12h, remove the gel, silver stain, and finally digital photography.
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2 test results
2.1 PCR amplification results
Figure 1 PCR amplification results of MyoG exon1-1 (1) (bp)
Figure 2 PCR amplification results of MyoG exon1-1(2) (bp)
2.2 PCR-SSCP test results
Figure 3 MyoG SSCP test results
In Fig. 3, SSCP detection results of PCR amplification products: 4, 6, 7, and 10 are AA genotypes; 1, 2, 5, and 8 are AB genotypes; and 3 and 9 are BB genotypes.
2.3 Results statistics
It can be seen from Table 1 that the effect of the MyoG gene polymorphism on the dripping rate of the pouch pigs did not reach a significant effect (P>0.05).
The effect on tenderness reached a significant level (P < 0.05). The AA type gene has the highest tenderness value, the AB type gene has the tenderness, and the BB type gene has the tenderness between the two.
There was no significant level of influence on the marble pattern (P>0.05). The effect of intramuscular fat reached a significant level (P<0.05). The content of the AB-type gene in the poached pork was higher than that of the AA-type gene and the BB-type gene by 0.16% and 0.29%, respectively.
The effect of pH on the pH value of MyoG gene did not reach a significant level at 45 min (P>0.05). However, the effect on the pH at 24 h achieved a significant effect (P < 0.05), but both were within the normal range. Table 1 Effect of polymorphism of MyoG gene on poultry pork
project | AA | AB | BB |
Drip rate /% | 1.38±0.41a | 1.27±0.37a | 1.34±0.08a |
Note: Different lowercase letters on the shoulders indicate significant differences (P<0.05), and the same letters indicate that the difference is not significant (P>0.05). The same below.
From the analysis of Table 2, it can be concluded that in the fiber area, fiber diameter and muscle fiber density, the AA type gene, the AB type gene and the BB type gene of the poached pork 3 reached a significant level (P < 0.05).
Table 2 Effect of MyoG gene polymorphism on the histological structure of pig muscle fibers
project | AA | AB | BB |
| Fiber area / μm2 Fiber diameter / μm Muscle fiber density / (root / mm2) | 20549±115.4c 30.27±0.19a 288.43±3.38b | 21351±97.83b 32.15±0.39a 294.47±8.11ab | 22090±103.22a 30.14±0.77b 308.59±10.02a |
3 Discussion
3.1 Polymorphism of MyoG gene
In this experiment, the pouch and the Changbai pig were selected as the target, and the MyoG gene polymorphism analysis was performed by PCR-SSCP. The result showed polymorphism in the exon I fragment. The results were compared with Soumillion et al. (1997) by PCR-RFLP method. MspI was digested and a total of 3 sites were found to be identical.
3.2 Effect of MyoG gene on pork quality index
The drip rate is an indicator of the hydraulic power of the pork. It is a measure of the loss of liquid without any external force. It can be seen from the test results that the dripping rate of the AB-type gene-packed pigs is significantly lower than that of the other two types, which indicates that the AB-type gene-loaded pork has a strong water-holding capacity. At present, research on pork tenderness has found that varieties and genetic factors have a great influence on the tenderness of meat. China's local pig breeds have good meat quality and fine muscle fibers. The muscle fat content is generally above 3%, even at 6% to 7%. The meat is tender and juicy, and the exotic varieties have been intramuscular due to excessive selection of lean meat rate. The fat content is reduced by 1% to 1.5%, which is far below the optimum range, so the meat is rough. The results of this experiment also proved that the tenderness of the local pig breed poached pigs was better than that of the imported pig breeds, and they all reached significant levels. The most tender ones are AB type pigs. The content of intramuscular fat directly affects the tenderness and marble of meat. The highest content of intramuscular fat purse pigs is AB type, which indicates that different genes play different roles in pig breeds. MyoG gene in marble. The effect of polymorphism on poached pork did not reach a significant level.
The pH is used as a criterion for evaluating inferior meat (Zeng Yongqing et al., 1990). The polymorphism of the MyoG gene had little effect on the pH of the meat at 45 min, and the pouch pig did not reach a significant level. However, the effect on the pH at 24 h reached a significant level, but both were within the normal range. Therefore, it is indicated that the three genes of the MyoG gene have little effect on the pH value.
According to some previous studies, the finer the muscle fibers, the more tender the meat is (Luo Jun, 1989). Many studies have shown that the diameter of muscle fibers of local pig breeds in China is thinner than that of imported pig breeds. Therefore, the tenderness of meat of local pig breeds is better than that of imported pig breeds. (Yang Bohui et al., 2001) studied the muscle fiber diameters of Jiulong Yak and Datong Yak, respectively, and found that the diameter of muscle fibers is related to the tenderness of meat.
3.3 Effect of MyoG gene on the histological structure of pork muscle fiber
In this experiment, the largest fiber area in the poached pork was BB type, and the three reached a significant level; the fiber diameter was the largest in AB type, and the three reached a significant level; the most dense fiber fiber was the BB type pouch pig. The three people also reached a significant level, and the fiber diameter and fiber area of ​​the poached pork were smaller than that of the Changbai pig, which further indicated that the tenderness of the poached pig was better than that of the imported pig.
(Reference)
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