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SNP-based analysis of genetic diversity reveals important alleles associated with seed size in rice.

Tang W, Wu T, Ye J, Sun J, Jiang Y, Yu J, Tang J, Chen G, Wang C, Wan J - BMC Plant Biol. (2016)

Bottom Line: Meanwhile, we identified polymorphic SNPs with large effects on protein-coding and miRNA genes.To validate the effect of the polymorphic SNPs, we further investigated a SNP (chr4:28,894,757) at the miRNA binding site in the 3'-UTR region of the locus Os4g48460, which is associated with rice seed size.Our study has identified the genome-wide SNPs by GBS of the parental varieties of RIL populations and identified CYP704A3, a miRNA-regulated gene that is responsible for rice seed length.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Crop Genetics and Germplasm Enhancement, Nanjing Agricultural University, 210095, Nanjing, China.

ABSTRACT

Background: Single-nucleotide polymorphisms (SNPs) have become the genetic markers of choice in various genetic, ecological, and evolutionary studies. Genotyping-by-sequencing (GBS) is a next-generation-sequencing based method that takes advantage of reduced representation to enable high-throughput genotyping using a large number of SNP markers.

Results: In the present study, the distribution of non-redundant SNPs in the parents of 12 rice recombination line populations was evaluated through GBS. A total of 45 Gigabites of nucleotide sequences conservatively provided satisfactory genotyping of rice SNPs. By assembling to the genomes of reference genomes of japonica Nipponbare, we detected 22,682 polymorphic SNPs that may be utilized for QTL/gene mapping with the Recombinant Inbred Lines (RIL) populations derived from these parental lines. Meanwhile, we identified polymorphic SNPs with large effects on protein-coding and miRNA genes. To validate the effect of the polymorphic SNPs, we further investigated a SNP (chr4:28,894,757) at the miRNA binding site in the 3'-UTR region of the locus Os4g48460, which is associated with rice seed size. Os4g48460 encodes a putative cytochrome P450, CYP704A3. Direct degradation of the 3'-UTR of the CYP704A3 gene by a miRNA (osa-miRf10422-akr) was validated by in planta mRNA degradation assay. We also showed that rice seeds of longer lengths may be produced by downregulating CYP704A3 via RNAi.

Conclusions: Our study has identified the genome-wide SNPs by GBS of the parental varieties of RIL populations and identified CYP704A3, a miRNA-regulated gene that is responsible for rice seed length.

No MeSH data available.


osa-miRf10422-akr precursors and its putative target gene. aCYP704A3 is a putative target of osa-miRf10422-akr. CYP704A3 structure and mutation sites are labeled as SNPs (blue) and the changed amino acid residues (red). b Significant differences in grain length between SNP genotypes of the CYP704A3 gene. **indicates a significance at P < 0.01. c qRT-PCR analysis shows the expression of CYP704A3 in maturing seeds. Asominori and IR24 were analyzed in terms of the expression of the CYP704A3 transcript. Seeds with longer lengths have lower levels of relative expression, similar to the other members of subfamily CYP450 such as CYP724B1, of which its mRNA accumulates at higher levels in CYP724B1 mutants with shorter seed compared to that in the wide-type. d Subcellular localization of the CYP704A3Protein. GFP signals of the CYP704A3-GFP fusion proteins localized in the endoplasmic reticulum of rice protoplasts
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Fig3: osa-miRf10422-akr precursors and its putative target gene. aCYP704A3 is a putative target of osa-miRf10422-akr. CYP704A3 structure and mutation sites are labeled as SNPs (blue) and the changed amino acid residues (red). b Significant differences in grain length between SNP genotypes of the CYP704A3 gene. **indicates a significance at P < 0.01. c qRT-PCR analysis shows the expression of CYP704A3 in maturing seeds. Asominori and IR24 were analyzed in terms of the expression of the CYP704A3 transcript. Seeds with longer lengths have lower levels of relative expression, similar to the other members of subfamily CYP450 such as CYP724B1, of which its mRNA accumulates at higher levels in CYP724B1 mutants with shorter seed compared to that in the wide-type. d Subcellular localization of the CYP704A3Protein. GFP signals of the CYP704A3-GFP fusion proteins localized in the endoplasmic reticulum of rice protoplasts

Mentions: We sequenced this gene that exhibited a total of four SNPs, with two present in the exon and two within the 3′-UTR. A G/A SNP (chr4:28,894,757) in the 3′-UTR was detected in the binding region between CYP704A3 and its miRNA gene, osa-miRf10422-akr (Fig. 3a).Fig. 3


SNP-based analysis of genetic diversity reveals important alleles associated with seed size in rice.

Tang W, Wu T, Ye J, Sun J, Jiang Y, Yu J, Tang J, Chen G, Wang C, Wan J - BMC Plant Biol. (2016)

osa-miRf10422-akr precursors and its putative target gene. aCYP704A3 is a putative target of osa-miRf10422-akr. CYP704A3 structure and mutation sites are labeled as SNPs (blue) and the changed amino acid residues (red). b Significant differences in grain length between SNP genotypes of the CYP704A3 gene. **indicates a significance at P < 0.01. c qRT-PCR analysis shows the expression of CYP704A3 in maturing seeds. Asominori and IR24 were analyzed in terms of the expression of the CYP704A3 transcript. Seeds with longer lengths have lower levels of relative expression, similar to the other members of subfamily CYP450 such as CYP724B1, of which its mRNA accumulates at higher levels in CYP724B1 mutants with shorter seed compared to that in the wide-type. d Subcellular localization of the CYP704A3Protein. GFP signals of the CYP704A3-GFP fusion proteins localized in the endoplasmic reticulum of rice protoplasts
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4837510&req=5

Fig3: osa-miRf10422-akr precursors and its putative target gene. aCYP704A3 is a putative target of osa-miRf10422-akr. CYP704A3 structure and mutation sites are labeled as SNPs (blue) and the changed amino acid residues (red). b Significant differences in grain length between SNP genotypes of the CYP704A3 gene. **indicates a significance at P < 0.01. c qRT-PCR analysis shows the expression of CYP704A3 in maturing seeds. Asominori and IR24 were analyzed in terms of the expression of the CYP704A3 transcript. Seeds with longer lengths have lower levels of relative expression, similar to the other members of subfamily CYP450 such as CYP724B1, of which its mRNA accumulates at higher levels in CYP724B1 mutants with shorter seed compared to that in the wide-type. d Subcellular localization of the CYP704A3Protein. GFP signals of the CYP704A3-GFP fusion proteins localized in the endoplasmic reticulum of rice protoplasts
Mentions: We sequenced this gene that exhibited a total of four SNPs, with two present in the exon and two within the 3′-UTR. A G/A SNP (chr4:28,894,757) in the 3′-UTR was detected in the binding region between CYP704A3 and its miRNA gene, osa-miRf10422-akr (Fig. 3a).Fig. 3

Bottom Line: Meanwhile, we identified polymorphic SNPs with large effects on protein-coding and miRNA genes.To validate the effect of the polymorphic SNPs, we further investigated a SNP (chr4:28,894,757) at the miRNA binding site in the 3'-UTR region of the locus Os4g48460, which is associated with rice seed size.Our study has identified the genome-wide SNPs by GBS of the parental varieties of RIL populations and identified CYP704A3, a miRNA-regulated gene that is responsible for rice seed length.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Crop Genetics and Germplasm Enhancement, Nanjing Agricultural University, 210095, Nanjing, China.

ABSTRACT

Background: Single-nucleotide polymorphisms (SNPs) have become the genetic markers of choice in various genetic, ecological, and evolutionary studies. Genotyping-by-sequencing (GBS) is a next-generation-sequencing based method that takes advantage of reduced representation to enable high-throughput genotyping using a large number of SNP markers.

Results: In the present study, the distribution of non-redundant SNPs in the parents of 12 rice recombination line populations was evaluated through GBS. A total of 45 Gigabites of nucleotide sequences conservatively provided satisfactory genotyping of rice SNPs. By assembling to the genomes of reference genomes of japonica Nipponbare, we detected 22,682 polymorphic SNPs that may be utilized for QTL/gene mapping with the Recombinant Inbred Lines (RIL) populations derived from these parental lines. Meanwhile, we identified polymorphic SNPs with large effects on protein-coding and miRNA genes. To validate the effect of the polymorphic SNPs, we further investigated a SNP (chr4:28,894,757) at the miRNA binding site in the 3'-UTR region of the locus Os4g48460, which is associated with rice seed size. Os4g48460 encodes a putative cytochrome P450, CYP704A3. Direct degradation of the 3'-UTR of the CYP704A3 gene by a miRNA (osa-miRf10422-akr) was validated by in planta mRNA degradation assay. We also showed that rice seeds of longer lengths may be produced by downregulating CYP704A3 via RNAi.

Conclusions: Our study has identified the genome-wide SNPs by GBS of the parental varieties of RIL populations and identified CYP704A3, a miRNA-regulated gene that is responsible for rice seed length.

No MeSH data available.