Exploiting cis-limited antigens in livestock

在畜牧业中，预先选择后代性别的能力将非常重要。在许多农场物种中，如猪或奶牛，雌性后代在经济上比雄性重要得多，而在其他物种，如赛马或肉牛，情况正好相反。特别值得一提的是，成年公猪的肉可能会有一种被称为野猪污染的味道，这意味着将完整的公猪饲养到高体重是不划算的。目前，通过提前屠宰或阉割公猪来避免公猪受到污染，这在伦理上是不可取的，在经济上也是浪费的。我们正在试图发现在受精之前对精子进行分类的方法，即区分携带X染色体的精子(将产生雌性后代)和携带Y染色体的精子(将产生雄性后代)。目前，做到这一点的唯一方法是使用染料对精子头部的DNA进行染色，并将DNA较多的精子与DNA较少的精子分开--这是可行的，因为X染色体略大于Y染色体。这种“流动分选”过程缓慢而繁琐，对猪等一些物种来说并不具有商业可行性。此外，流动分选过程涉及对DNA进行染色并向其照射紫外线激光，这两种操作都可能损害DNA。因此，迫切需要更有效、更安全的方法来分离X和Y精子。我们正在使用两种方法来解决这个问题。首先，我们正在研究X和Y精子是否自然地在其表面携带标记，从而区分它们。我们将通过使用流动分选来分离发育中的X-和Y-精子，并寻找在每种细胞类型中哪些基因是活跃的。如果我们发现只在两种细胞类型中的一种中打开的基因，这些基因很可能在开发分离这两种细胞的方法时有用。我们将在正常小鼠、自然产生60%雌性后代的小鼠品系以及改变了我们认为涉及的基因水平的转基因品系中进行这一研究。从以前的工作中，我们知道，在这些性别比例扭曲的菌株中，产生了相同数量的X-和Y-精子，但Y精子在使卵子受精方面效率较低。这意味着这些菌株将帮助我们定位感兴趣的基因--即使X和Y精子彼此不同的基因。其次，我们想测试是否可以使用人造表面标记来分离精子。为了做到这一点，我们将培育在其精子细胞中表达标记蛋白的转基因小鼠。理想情况下，含有转基因的精子细胞将在细胞表面显示这个“标签”，而那些不含转基因的精子细胞将保持不加标签。这将使我们能够分离这两种类型的精子，并只使用未标记的精子来繁殖下一代。在家畜中，这将使精子选择成为可能，同时防止转基因生物进入食物链。

The ability to pre-select the sex of offspring would be hugely important in animal husbandry. In many farm species such as pigs or dairy cattle, female offspring are much more economically important than males, while in other species such as racehorses or beef cattle, the opposite situation holds. In particular, meat from adult male boars can have an "off" flavour known as boar taint, meaning it is uneconomical to raise intact male pigs to high weights. Currently, boar taint is avoided by early slaughter or castration of male pigs, which is ethically undesirable as well as economically wasteful. We are trying to discover ways to sort sperm before fertilisation occurs, i.e. to distinguish between sperm bearing an X chromosome (which will give female offspring) and those bearing a Y chromosome (which will give males). Currently the only way to do this is by using a dye to stain the DNA in the sperm head, and separating the sperm that have more DNA from those that have less - this works because the X chromosome is slightly larger than the Y. This "flow-sorting" process is slow and cumbersome and not commercially viable for some species such as pig. Also, the flow-sorting process involves staining the DNA and shining an ultraviolet laser on it, both of which can potentially damage the DNA. There is thus a strong need for more efficient and safer ways of separating X and Y sperm. We are using two approaches to tackle the problem.Firstly, we are looking to see whether X and Y sperm naturally carry markers on their surface allowing them to be distinguished. We will do this by using flow-sorting to separate developing X- and Y-bearing sperm and looking to see which genes are active in each cell type. If we find genes which are only switched on in one of the two cell types, these are likely to be be useful in developing ways to separate the two. We will carry out this search in normal mice, in a strain of mice which naturally produce 60% female offspring, and in a transgenic strain which has altered levels of a gene we believe to be involved. From previous work, we know that in these strains with a skewed sex ratio, equal numbers of X- and Y-bearing sperm are produced, but that the Y sperm are less effective at fertilising the egg. This means that these strains will help us to home in on genes of interest - i.e. the ones which make X and Y sperm different from each other. Secondly, we want to test whether we can separate sperm using an artificial surface marker. To do this, we will breed transgenic mice which express a marker protein in their sperm cells. Ideally, sperm cells containg the transgene will show this "tag" on the cell surface, while those which do not contain the transgene will remain untagged. This will then allow us to separate the two types of sperm and only use untagged sperm to breed the next generation. In livestock, this would enable sperm selection while preventing genetically modified organisms entering the food chain.