Rothamsted Research, Harpenden, Hertfordshire, UK.
Earlham Institute, Norwich Research Park, Norwich, UK.
Ann Bot. 2021 Sep 7;128(5):627-637. doi: 10.1093/aob/mcab101.
The C4Urochloa species (syn. Brachiaria) and Megathyrsus maximus (syn. Panicum maximum) are used as pasture for cattle across vast areas in tropical agriculture systems in Africa and South America. A key target for variety improvement is forage quality: enhanced digestibility could decrease the amount of land required per unit production, and enhanced lipid content could decrease methane emissions from cattle. For these traits, loss-of-function (LOF) alleles in known gene targets are predicted to improve them, making a reverse genetics approach of allele mining feasible. We therefore set out to look for such alleles in diverse accessions of Urochloa species and Megathyrsus maximus from the genebank collection held at the CIAT.
We studied allelic diversity of 20 target genes (11 for digestibility, nine for lipid content) in 104 accessions selected to represent genetic diversity and ploidy levels of U. brizantha, U. decumbens, U. humidicola, U. ruziziensis and M. maximum. We used RNA sequencing and then bait capture DNA sequencing to improve gene models in a U. ruziziensis reference genome to assign polymorphisms with high confidence.
We found 953 non-synonymous polymorphisms across all genes and accessions; within these, we identified seven putative LOF alleles with high confidence, including those in the non-redundant SDP1 and BAHD01 genes present in diploid and tetraploid accessions. These LOF alleles could respectively confer increased lipid content and digestibility if incorporated into a breeding programme.
We demonstrated a novel, effective approach to allele discovery in diverse accessions using a draft reference genome from a single species. We used this to find gene variants in a collection of tropical grasses that could help reduce the environmental impact of cattle production.
C4 雀稗属（syn. 象草属）和大黍（syn. 象草属）被广泛用于非洲和南美洲热带农业系统的牛类牧场。品种改良的一个关键目标是饲草质量：提高消化率可以减少每单位产量所需的土地量，而提高脂质含量可以减少牛类的甲烷排放量。对于这些性状，已知基因靶标中的功能丧失（LOF）等位基因预计会改善它们，从而使等位基因挖掘的反向遗传学方法成为可行。因此，我们着手在 CIAT 基因库中收集的来自 Urochloa 属和大黍属的不同品种中寻找这些等位基因。
我们研究了 104 个品种中 20 个目标基因（11 个用于消化率，9 个用于脂质含量）的等位基因多样性，这些品种选择代表了 U. brizantha、U. decumbens、U. humidicola、U. ruziziensis 和 M. maximum 的遗传多样性和倍性水平。我们使用 RNA 测序，然后使用诱饵捕获 DNA 测序来改进 U. ruziziensis 参考基因组中的基因模型，以高度置信度分配多态性。
我们在所有基因和品种中发现了 953 个非同义多态性；在这些多态性中，我们确定了七个具有高度置信度的推定 LOF 等位基因，包括存在于二倍体和四倍体品种中的非冗余 SDP1 和 BAHD01 基因中的那些。如果将这些 LOF 等位基因纳入一个育种计划，它们可以分别赋予更高的脂质含量和消化率。
我们展示了一种在使用单一物种的草案参考基因组的多种品种中发现等位基因的新颖、有效的方法。我们利用这种方法在热带牧草的收集物中发现了可能有助于减少牛类生产对环境影响的基因变体。
