Abstract
Key message
Insertion of a solo LTR, which possesses strong bidirectional, stem-specific promoter activities, is associated with the evolution of a dwarfing apple spur mutation.
Abstract
Spur mutations in apple scions revolutionized global apple production. Since long terminal repeat (LTR) retrotransposons are tightly related to natural mutations, inter-retrotransposon-amplified polymorphism technique and genome walking were used to find sequences in the apple genome based on these LTRs. In ‘Red Delicious’ spur mutants, a novel, 2190-bp insertion was identified as a spur-specific, solo LTR (sLTR) located at the 1038th nucleotide of another sLTR, which was 1536 bp in length. This insertion-within-an-insertion was localized within a preexisting Gypsy-50 retrotransposon at position 3,762,767 on chromosome 4. The analysis of transcriptional activity of the two sLTRs (the 2190- and 1536-bp inserts) indicated that the 2190-bp sLTR is a promoter, capable of bidirectional transcription. GUS expression in the 2190-bp-sense and 2190-bp-antisense transgenic lines was prominent in stems. In contrast, no promoter activity from either the sense or the antisense strand of the 1536-bp sLTR was detected. From ~150 kb of DNA on each side of the 2190 bp, sLTR insertion site, corresponding to 300 kb of the ‘Golden Delicious’ genome, 23 genes were predicted. Ten genes had predicted functions that could affect shoot development. This first report, of a sLTR insertion associated with the evolution of apple spur mutation, will facilitate apple breeding, cloning of spur-related genes, and discovery of mechanisms behind dwarf habit.
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Abbreviations
- ACX1:
-
Acyl-coenzyme A oxidase 1
- ACO:
-
1-Aminocyclopropane-1-carboxylate oxidase
- CIPK:
-
CBL-interacting protein kinase
- ERF:
-
Ethylene-responsive transcription factor
- IRAP:
-
Inter-retrotransposon amplified polymorphism
- LTR:
-
Long terminal repeat
- LRNs:
-
LTR retrotransposons
- ORFs:
-
Open reading frames
- QTL:
-
Quantitative trait loci
- IPCR:
-
Reverse PCR
- S-SAP:
-
Sequence-specific amplification polymorphism
- sLTR:
-
Solo LTR
- TAIL-PCR:
-
Thermal asymmetric interlaced PCR
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Acknowledgements
This work was supported by two Grants-in-Aid for Scientific Research from the State National Natural Science Foundation of China (Nos. 31372043 and 31071776).
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Communicated by Dr. Prakash P. Kumar.
Electronic supplementary material
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299_2017_2160_MOESM1_ESM.pdf
Supplementary material 1 (PDF 105 kb) The schematic of IRAP and genome walking in ‘Red Delicious’ and its two spur mutants, ‘Chinese Marshal 1’ and ‘Oregon spur Delicious’. Red box represents Fragment-1 from the two spur mutants amplified using IRAP technique. Green box represents Sequence-2 from the two spur mutants amplified using TAIL-PCR technique. Yellow box represents Sequence-3 amplified from the two spur mutants using IPCR. The primers LP-1 and LP-2 were designed based on the sequence of the apple genome to amplify the sequence flanking Sequence-1, 2 and 3
299_2017_2160_MOESM2_ESM.pdf
Supplementary material 2 (PDF 65 kb) The target site duplication (TSD, red) and inverted repeat (IR, italics) of the 1536-bp sLTR insertion (green) are shown. The TSD and IR of the 2190-bp sLTR insertion (yellow) are boxed and underlined, respectively. The TSD and IR of Gypsy-50 are indicated in red and double underlined. LTRs of Gypsy-50 are indicated in blue. The two ‘CGGGG’ sites are shaded in blue
299_2017_2160_MOESM3_ESM.png
Supplementary material 3 (PNG 4740 kb) The alignment of the 122 LTR-like copies found in apple using the 2190-bp fragment, ranging in size from 1777-bp to 2190-bp
299_2017_2160_MOESM4_ESM.pdf
Supplementary material 4 (PDF 86 kb) Primers used for overlapped amplification of products of PL-1 and PL-2, amplified by primers LP-1 and LP-2
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Han, M., Sun, Q., Zhou, J. et al. Insertion of a solo LTR retrotransposon associates with spur mutations in ‘Red Delicious’ apple (Malus × domestica). Plant Cell Rep 36, 1375–1385 (2017). https://doi.org/10.1007/s00299-017-2160-x
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DOI: https://doi.org/10.1007/s00299-017-2160-x