Universidade Federal do Recôncavo da Bahia, Campus Cruz das Almas, CEP, Cruz das Almas, BA, 44380-000, Brazil.
International Center for Tropical Agriculture (CIAT), A.A 6713, Cali, Colombia.
BMC Plant Biol. 2020 Apr 15;20(1):164. doi: 10.1186/s12870-020-02379-3.
The granule-bound starch synthase I (GBSSI) enzyme is responsible for the synthesis of amylose, and therefore, its absence results in individuals with a waxy starch phenotype in various amylaceous crops. The validation of mutation points previously associated with the waxy starch phenotype in cassava, as well as the identification of alternative mutant alleles in the GBSSI gene, can allow the development of molecular-assisted selection to introgress the waxy starch mutation into cassava breeding populations.
A waxy cassava allele has been identified previously, associated with several SNPs. A particular SNP (intron 11) was used to develop SNAP markers for screening heterozygote types in cassava germplasm. Although the molecular segregation corresponds to the expected segregation at 3:1 ratio (dominant gene for the presence of amylose), the homozygotes containing the SNP associated with the waxy mutation did not show waxy phenotypes. To identify more markers, we sequenced the GBSS gene from 89 genotypes, including some that were segregated from a cross with a line carrying the known waxy allele. As a result, 17 mutations in the GBSSI gene were identified, in which only the deletion in exon 6 (MeWxEx6-del-C) was correlated with the waxy phenotype. The evaluation of mutation points by discriminant analysis of principal component analysis (DAPC) also did not completely discriminate the waxy individuals. Therefore, we developed Kompetitive Allele Specific PCR (KASP) markers that allowed discrimination between WX and wx alleles. The results demonstrated the non-existence of heterozygous individuals of the MeWxEx6-del-C deletion in the analyzed germplasm. Therefore, the deletion MeWxEx6-del-C should not be used for assisted selection in genetic backgrounds different from the original source of waxy starch. Also, the alternative SNPs identified in this study were not associated with the waxy phenotype when compared to a panel of accessions with high genetic diversity.
Although the GBSSI gene can exhibit several mutations in cassava, only the deletion in exon 6 (MeWxEx6-del-C) was correlated with the waxy phenotype in the original AM206-5 source.
颗粒结合淀粉合成酶 I（GBSSI）酶负责直链淀粉的合成，因此，其缺失导致各种含淀粉作物的蜡质淀粉表型个体。验证先前与木薯蜡质淀粉表型相关的突变点，并在 GBSSI 基因中鉴定替代的突变等位基因，可允许开发分子辅助选择，将蜡质淀粉突变体导入木薯育种群体。
先前已鉴定出一种与多个 SNP 相关的蜡质木薯等位基因。一个特定的 SNP（内含子 11）被用于开发 SNAP 标记，以筛选木薯种质中的杂合体型。尽管分子分离与预期的 3:1 分离比（存在直链淀粉的显性基因）相对应，但含有与蜡质突变相关 SNP 的纯合子并未表现出蜡质表型。为了鉴定更多的标记，我们对包括来自已知携带蜡质等位基因的杂交种分离的 89 个基因型的 GBSS 基因进行了测序。结果，在 GBSSI 基因中发现了 17 个突变，其中只有外显子 6 的缺失（MeWxEx6-del-C）与蜡质表型相关。通过主成分分析判别分析（DAPC）对突变点的评估也不能完全区分蜡质个体。因此，我们开发了 Kompetitive Allele Specific PCR（KASP）标记，能够区分 WX 和 wx 等位基因。结果表明，在所分析的种质中，不存在 MeWxEx6-del-C 缺失的杂合个体。因此，MeWxEx6-del-C 的缺失不应在与蜡质淀粉原始来源不同的遗传背景下用于辅助选择。此外，与具有高度遗传多样性的一组访问者相比，本研究中鉴定的替代 SNP 与蜡质表型无关。
尽管 GBSSI 基因在木薯中可能表现出多种突变，但只有外显子 6 的缺失（MeWxEx6-del-C）与 AM206-5 原始来源的蜡质表型相关。
