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Identification of Genes Potentially Associated with the Fertility Instability of S-Type Cytoplasmic Male Sterility in Maize via Bulked Segregant RNA-Seq

View Article: PubMed Central - PubMed

ABSTRACT

S-type cytoplasmic male sterility (CMS-S) is the largest group among the three major types of CMS in maize. CMS-S exhibits fertility instability as a partial fertility restoration in a specific nuclear genetic background, which impedes its commercial application in hybrid breeding programs. The fertility instability phenomenon of CMS-S is controlled by several minor quantitative trait locus (QTLs), but not the major nuclear fertility restorer (Rf3). However, the gene mapping of these minor QTLs and the molecular mechanism of the genetic modifications are still unclear. Using completely sterile and partially rescued plants of fertility instable line (FIL)-B, we performed bulk segregant RNA-Seq and identified six potential associated genes in minor effect QTLs contributing to fertility instability. Analyses demonstrate that these potential associated genes may be involved in biological processes, such as floral organ differentiation and development regulation, energy metabolism and carbohydrates biosynthesis, which results in a partial anther exsertion and pollen fertility restoration in the partially rescued plants. The single nucleotide polymorphisms (SNPs) identified in two potential associated genes were validated to be related to the fertility restoration phenotype by KASP marker assays. This novel knowledge contributes to the understanding of the molecular mechanism of the partial fertility restoration of CMS-S in maize and thus helps to guide the breeding programs.

No MeSH data available.


Identification of genomic regions contributing to the fertility instability in FIL-B by BSR-Seq using the Euclidean distance (ED) algorithm.A. The ED scores raised to the fifth power across the genome. Each dot represents each SNP identified from the RNA-Seq, and the different colors designate the different chromosomes as indicated on the X-axis. For all of the panels, the gray vertical dotted lines delineate the chromosome edges, and the width of the chromosome represents the relative numbers of SNPs identified. The pink horizontal dotted lines represent the significant threshold of the 99% percentile of the ED5. B. The ED5 scores of a close-up of chromosome 2. C. The Loess fit curve calculated from A. D. The Loess fit curve of a close-up of chromosome 2 with the physical position indicated on X-axis. Each peak represents a possible associated genomic region.
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pone.0163489.g004: Identification of genomic regions contributing to the fertility instability in FIL-B by BSR-Seq using the Euclidean distance (ED) algorithm.A. The ED scores raised to the fifth power across the genome. Each dot represents each SNP identified from the RNA-Seq, and the different colors designate the different chromosomes as indicated on the X-axis. For all of the panels, the gray vertical dotted lines delineate the chromosome edges, and the width of the chromosome represents the relative numbers of SNPs identified. The pink horizontal dotted lines represent the significant threshold of the 99% percentile of the ED5. B. The ED5 scores of a close-up of chromosome 2. C. The Loess fit curve calculated from A. D. The Loess fit curve of a close-up of chromosome 2 with the physical position indicated on X-axis. Each peak represents a possible associated genomic region.

Mentions: The sequencing reads were aligned to the B73 reference genome to obtain the SNPs in both of the bulks. A total of 171,594 SNPs and 170,820 SNPs were identified in bulk F and bulk S, respectively. To map the QTLs controlling fertility restoration, the Euclidean Distance (ED) algorithm was applied to measure allele segregation and to identify the linked genomic loci based on the SNPs between the two bulks. A statistically significant peak of ED was observed on chromosome 2 (Fig 4A and 4B), indicating its association to fertility instability in FIL-B. Seven genomic regions were associated with fertility instability by using the 99% percentile of the ED5 (0.17678) as the significant cutoff (Fig 4C and 4D), among which two regions consisted of only a couple of bases. The other 5 regions were all located on the short arm of chromosome 2 (Table 2). To identify the potential associated genes within these 5 genetic loci, the annotated genes were further screened based on their expression pattern within the two bulks. A total of 12 genes (Table 3) exhibited differential expression, 8 genes were located in region 2 and 4 genes were located in region 5, with no genes identified in the other 3 regions (Table 2). Of these genes, GRMZM2G315401 and GRMZM2G434669 encode the serine/arginine -rich protein 45, GRMZM2G430362 encodes an ATP-dependent RNA helicase SUV3, GRMZM2G474783 and GRMZM2G010338 encode the leucine-rich repeat extensin-like protein 5 and a putative SPRY-domain family protein, respectively, and GRMZM2G127173 encodes an uncoupling protein regulating ATP biosynthesis. The 6 above mentioned genes were selected as potential associated genes for the following expression study because of their possible function as regulatory genes. Moreover, GRMZM2G012328 is predicted as pectinesterase, GRMZM2G145758 possibly encodes histone H3, and AC191050.3_FG003 encodes a potential precursor of the globulin-1 S allele, all of which may involve in cell wall metabolism and modification. The other three genes were annotated as hypothetical proteins and thus need further investigation.


Identification of Genes Potentially Associated with the Fertility Instability of S-Type Cytoplasmic Male Sterility in Maize via Bulked Segregant RNA-Seq
Identification of genomic regions contributing to the fertility instability in FIL-B by BSR-Seq using the Euclidean distance (ED) algorithm.A. The ED scores raised to the fifth power across the genome. Each dot represents each SNP identified from the RNA-Seq, and the different colors designate the different chromosomes as indicated on the X-axis. For all of the panels, the gray vertical dotted lines delineate the chromosome edges, and the width of the chromosome represents the relative numbers of SNPs identified. The pink horizontal dotted lines represent the significant threshold of the 99% percentile of the ED5. B. The ED5 scores of a close-up of chromosome 2. C. The Loess fit curve calculated from A. D. The Loess fit curve of a close-up of chromosome 2 with the physical position indicated on X-axis. Each peak represents a possible associated genomic region.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5036866&req=5

pone.0163489.g004: Identification of genomic regions contributing to the fertility instability in FIL-B by BSR-Seq using the Euclidean distance (ED) algorithm.A. The ED scores raised to the fifth power across the genome. Each dot represents each SNP identified from the RNA-Seq, and the different colors designate the different chromosomes as indicated on the X-axis. For all of the panels, the gray vertical dotted lines delineate the chromosome edges, and the width of the chromosome represents the relative numbers of SNPs identified. The pink horizontal dotted lines represent the significant threshold of the 99% percentile of the ED5. B. The ED5 scores of a close-up of chromosome 2. C. The Loess fit curve calculated from A. D. The Loess fit curve of a close-up of chromosome 2 with the physical position indicated on X-axis. Each peak represents a possible associated genomic region.
Mentions: The sequencing reads were aligned to the B73 reference genome to obtain the SNPs in both of the bulks. A total of 171,594 SNPs and 170,820 SNPs were identified in bulk F and bulk S, respectively. To map the QTLs controlling fertility restoration, the Euclidean Distance (ED) algorithm was applied to measure allele segregation and to identify the linked genomic loci based on the SNPs between the two bulks. A statistically significant peak of ED was observed on chromosome 2 (Fig 4A and 4B), indicating its association to fertility instability in FIL-B. Seven genomic regions were associated with fertility instability by using the 99% percentile of the ED5 (0.17678) as the significant cutoff (Fig 4C and 4D), among which two regions consisted of only a couple of bases. The other 5 regions were all located on the short arm of chromosome 2 (Table 2). To identify the potential associated genes within these 5 genetic loci, the annotated genes were further screened based on their expression pattern within the two bulks. A total of 12 genes (Table 3) exhibited differential expression, 8 genes were located in region 2 and 4 genes were located in region 5, with no genes identified in the other 3 regions (Table 2). Of these genes, GRMZM2G315401 and GRMZM2G434669 encode the serine/arginine -rich protein 45, GRMZM2G430362 encodes an ATP-dependent RNA helicase SUV3, GRMZM2G474783 and GRMZM2G010338 encode the leucine-rich repeat extensin-like protein 5 and a putative SPRY-domain family protein, respectively, and GRMZM2G127173 encodes an uncoupling protein regulating ATP biosynthesis. The 6 above mentioned genes were selected as potential associated genes for the following expression study because of their possible function as regulatory genes. Moreover, GRMZM2G012328 is predicted as pectinesterase, GRMZM2G145758 possibly encodes histone H3, and AC191050.3_FG003 encodes a potential precursor of the globulin-1 S allele, all of which may involve in cell wall metabolism and modification. The other three genes were annotated as hypothetical proteins and thus need further investigation.

View Article: PubMed Central - PubMed

ABSTRACT

S-type cytoplasmic male sterility (CMS-S) is the largest group among the three major types of CMS in maize. CMS-S exhibits fertility instability as a partial fertility restoration in a specific nuclear genetic background, which impedes its commercial application in hybrid breeding programs. The fertility instability phenomenon of CMS-S is controlled by several minor quantitative trait locus (QTLs), but not the major nuclear fertility restorer (Rf3). However, the gene mapping of these minor QTLs and the molecular mechanism of the genetic modifications are still unclear. Using completely sterile and partially rescued plants of fertility instable line (FIL)-B, we performed bulk segregant RNA-Seq and identified six potential associated genes in minor effect QTLs contributing to fertility instability. Analyses demonstrate that these potential associated genes may be involved in biological processes, such as floral organ differentiation and development regulation, energy metabolism and carbohydrates biosynthesis, which results in a partial anther exsertion and pollen fertility restoration in the partially rescued plants. The single nucleotide polymorphisms (SNPs) identified in two potential associated genes were validated to be related to the fertility restoration phenotype by KASP marker assays. This novel knowledge contributes to the understanding of the molecular mechanism of the partial fertility restoration of CMS-S in maize and thus helps to guide the breeding programs.

No MeSH data available.