F1代转CuZnSOD基因猪的制备与研究

doi:10.11843/j.issn.0366-6964.2016.01.003

摘要/Abstract

摘要：

为进一步研究CuZnSOD基因对肉质表达调控的作用机理，探究通过磁性纳米颗粒介导精子载体法制备转基因猪的可行性。本试验在重组质粒先后与磁性纳米颗粒和精液共孵育的基础上，再通过人工授精获得F₁代仔猪。对繁殖得到的F₁代个体进行PCR、荧光定量PCR及Western blot检测，对屠宰的4头阳性猪采用绝对定量PCR进行转基因拷贝数鉴定，F₁代个体达100 kg左右时抽样屠宰进行肉质及抗氧化性能测定。结果表明，F₁代阳性率为30.23%。阳性组CuZnSOD在5种组织中的RNA和蛋白水平上的表达量较阴性组均上调。本试验得到的标准曲线：ΔCt=－3.121 6 lgN+17.281(R²=0.992 1，P<0.001)，4头阳性猪中外源基因拷贝数分别为25.89、80.61、106.69、83.03。肉质及抗氧化性能测定结果显示，阳性组肌肉滴水损失显著低于阴性组(P<0.05)；阳性组肌肉T-SOD_1d、T-SOD_2d活性显著高于阴性组，MDA_1d、MDA_2d含量显著低于阴性组(P<0.05)。研究结果表明：CuZnSOD基因的过表达可以提高肌肉的抗氧化性能，降低滴水损失，进而改善肉质，同时，证明磁性纳米颗粒介导的精子载体法可以用于制备转基因动物。

Abstract:

The aim of this study was to further research the mechanism of how CuZnSOD gene effect the meat quality by expression and regulation，and explore the possibility to generate transgenic pigs by sperm carrier method mediated by magnetic nanoparticles.F₁ generation individuals were obtained by artificial insemination on the basis of the recombinant vector incubated with magnetic nanoparticles and semen，respectively.F₁ individuals were detected by PCR，qRT-PCR and Western blot.Exogenous gene copy number in the 4 slaughtered transgenic positive pigs were estimated by absolute qPCR.Individuals from positive and negative groups were selected at the weight of about 100 kg and slaughtered to determine the meat quality and the antioxidant properties.The results showed that：The positive rate was 30.23% in F₁individuals.Compared with the negative group，the CuZnSOD expression at RNA and protein levels in 5 tissues were up-regulated in positive group.The standard curve in this study was：ΔCt=-3.121 6 lgN+17.281(R²=0.992 1，P<0.001).The exogenous gene copy number in the 4 positive pigs was 25.89，80.61，106.69 and 83.03，respectively.The results also showed that：Compared with the negative group，the positive group was significantly increased in muscle T-SOD_1dand T-SOD_2d activity(P<0.05)，but decreased in muscle drip loss，the content of MDA_1d and MDA_2d(P<0.05)，respectively.The results indicated that the overexpression of CuZnSOD gene could elevate the antioxidant properties，reduce the drip loss and further improve the meat quality.Meanwhile，our results proved that transgenic animals could be generated by sperm carrier method mediated by magnetic nanoparticles.

中图分类号:

S828
S813.3

王守栋，房国锋，曾勇庆，陈伟，李川皓，王延东. F₁代转CuZnSOD基因猪的制备与研究[J]. 畜牧兽医学报, 2016, 47(1): 16-24.

WANG Shou-dong，FANG Guo-feng，ZENG Yong-qing，CHEN Wei，LI Chuan-hao，WANG Yan-dong. Preparation and Study of CuZnSOD Gene Transgenic Pigs in F₁ Generation[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2016, 47(1): 16-24.

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F₁代转CuZnSOD基因猪的制备与研究

Preparation and Study of CuZnSOD Gene Transgenic Pigs in F₁ Generation

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