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QTL-Seq and Sequence Assembly Rapidly Mapped the Gene BrMYBL2.1 for the Purple Trait in Brassica rapa.

ABSTRACT: Anthocyanins have strong antioxidant activity and are believed to be healthy for human beings. The Brassica rapa L. ssp. chinensis var. purpurea "Zicaitai" is rich in anthocyanins. We constructed an F₂ population of Zicaitai and "Caixin" (Brassica rapa ssp. parachinensis) and it shows clear segregation of the purple phenotype (i.e., variation in anthocyanin enrichment). Here, quantitative trait locus (QTL)-Seq was performed with two sample groups from the F₂ population: one exhibiting an intense purple phenotype and the other showed a completely green phenotype. The results showed that the QTL-Seq and linkage analysis located different major loci. This indicates that there are two major genetic factors that plays different roles in regulating anthocyanin enrichment in Zicaitai. This was further supported by the data simulation of an in silico F₂ population that QTL-Seq and linkage analysis can locate different major loci. Furthermore, the draft genomes of the two parents (Zicaitai and Caixin) were assembled and utilized to search for mutations in candidate genes. A ~100-bp insertion was found in the third exon of gene BrMYBL2.1 in Zicaitai. BrMYBL2.1 is a negative regulator of anthocyanin biosynthesis, while BrEGL3.2-previously located by linkage mapping-is a positive regulator. For these populations with multiple genes contributing large effects to a trait, a strategy of low depth re-sequencing of F₂ individuals followed by QTL-Seq analysis with the free combination of sample groups is proposed. Furthermore, draft-sequence assembly of parental genomes together with QTL mapping is suggested as an efficient means for fine-mapping genes rapidly in segregating populations.

SUBMITTER: Zhang X

PROVIDER: S-EPMC7012920 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundBrassica rapa is an economically important crop and a model plant for studies concerning polyploidization and the evolution of extreme morphology. The multinational B. rapa Genome Sequencing Project (BrGSP) was launched in 2003. In 2008, next generation sequencing technology was used to sequence the B. rapa genome. Several maps concerning B. rapa pseudochromosome assembly have been published but their coverage of the genome is incomplete, anchoring approximately 73.6% of the scaffolds on to chromosomes. Therefore, a new genetic map to aid pseudochromosome assembly is required.ResultsThis study concerns the construction of a reference genetic linkage map for Brassica rapa, forming the backbone for anchoring sequence scaffolds of the B. rapa genome resulting from recent sequencing efforts. One hundred and nineteen doubled haploid (DH) lines derived from microspore cultures of an F1 cross between a Chinese cabbage (B. rapa ssp. pekinensis) DH line (Z16) and a rapid cycling inbred line (L144) were used to construct the linkage map. PCR-based insertion/deletion (InDel) markers were developed by re-sequencing the two parental lines. The map comprises a total of 507 markers including 415 InDels and 92 SSRs. Alignment and orientation using SSR markers in common with existing B. rapa linkage maps allowed ten linkage groups to be identified, designated A01-A10. The total length of the linkage map was 1234.2 cM, with an average distance of 2.43 cM between adjacent marker loci. The lengths of linkage groups ranged from 71.5 cM to 188.5 cM for A08 and A09, respectively. Using the developed linkage map, 152 scaffolds were anchored on to the chromosomes, encompassing more than 82.9% of the B. rapa genome. Taken together with the previously available linkage maps, 183 scaffolds were anchored on to the chromosomes and the total coverage of the genome was 88.9%.ConclusionsThe development of this linkage map is vital for the integration of genome sequences and genetic information, and provides a useful resource for the international Brassica research community.

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QTL-Seq and Sequence Assembly Rapidly Mapped the Gene BrMYBL2.1 for the Purple Trait in Brassica rapa.

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