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| Status |
Public on Nov 07, 2014 |
| Title |
Methylation Affects Transposition and Splicing of a Large CACTA Transposable Element From a MYB Transcription Factor Regulating Anthocyanin Synthase (ANS) Genes in Soybean Seed Coats (Bisulfite-Seq) |
| Organism |
Glycine max |
| Experiment type |
Methylation profiling by high throughput sequencing
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| Summary |
We determined the molecular basis of three soybean lines that vary in seed coat color at the R locus which is thought to encode a MYB transcription factor. RM55-rm is homozygous for a mutable allele (rm) that specifies black and brown striped seeds; RM30-R* is a stable black revertant isoline derived from the mutable line; and RM38-r has brown seed coats due to a recessive r allele shown to translate a truncated MYB protein. Using long range PCR, 454 sequencing of amplicons, and whole genome re-sequencing, we determined that the variegated RM55-rm line had a 13 kb CACTA subfamily transposon insertion (designated TgmR*) at a position 110 bp from the beginning of Intron2 of the R locus, Glyma09g36983. Although the MYB encoded by R was expressed at only very low levels in older seed coats of the black revertant RM30-R* line, it upregulated expression of anthocyanidin synthase genes (ANS2, ANS3) to promote the synthesis of anthocyanins. Surprisingly, the RM30-R* revertant also carried the 13 kb TgmR* insertion in Intron2. Using RNA-Seq, we showed that intron splicing was accurate, albeit at lower levels, despite the presence of the 13kb TgmR* element. As determined by whole genome methylation sequencing, we demonstrate that the TgmR* sequence was relatively more methylated in RM30-R* than in the mutable RM55-rm progenitor line. The stabilized and more methylated RM30-R* revertant line apparently lacks effective binding of a transposae to its subterminal repeats, thus allowing intron splicing to proceed resulting in sufficient MYB protein to stimulate anthocyanin production and thus black seed coats. In this regard, the TgmR* element in soybean resembles McClintock’s Spm-suppressible and change-of-state alleles of maize. This comparison explains the opposite effects of the TgmR* element on intron splicing of the MYB gene in which it resides depending on the methylation state of the element.
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| Overall design |
The genomic DNAs extracted from shoot tips of two seed color soybean lines, RM55-rm (black/brown striped) and RM30-R* (black) were sequenced in an Illumina HiSeq 2000 following the the Bisulfite (BS) sequencing method.
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| Contributor(s) |
Zabala G, Vodkin LO |
| Citation(s) |
25369033 |
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| Submission date |
Sep 04, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Lila O. Vodkin |
| E-mail(s) |
[email protected]
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| Phone |
217-244-6147
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| Organization name |
University of Illinois
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| Department |
Crop Sciences
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| Lab |
Lila Vodkin
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| Street address |
1201 W. Gregory Dr.
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| City |
Urbana |
| State/province |
IL |
| ZIP/Postal code |
61801 |
| Country |
USA |
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| Platforms (1) |
| GPL15008 |
Illumina HiSeq 2000 (Glycine max) |
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| Samples (2) |
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| This SubSeries is part of SuperSeries: |
| GSE61118 |
Methylation Affects Transposition and Splicing of a Large CACTA Transposable Element From a MYB Transcription Factor Regulating Anthocyanin Synthase (ANS) Genes in Soybean Seed Coats |
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| Relations |
| BioProject |
PRJNA260283 |
| SRA |
SRP046241 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data not provided for this record |
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