Abstract
Protocorm-like body (PLB) formation is an efficient method that has been widely used in the propagation of orchids. There is little information regarding the genome-wide molecular spectrum of somaclonal variation in PLB-regenerated orchids. We used specific-locus amplified fragment sequencing (SLAF-seq) to investigate the variations of single nucleotide polymorphisms (SNPs) and insertion-deletions (InDels) in somaclones of Oncidium ‘Milliongolds’, which was regenerated via PLB formation after in vitro culturing for 10 years. A high average variation rate (1.00 × 10− 3) was detected in 11 somaclones, and the most abundant variations were line specific. The average Ti/Tv ratio was 2.17, and G:C > C:G was the rarest type of SNP. The SNPs adjacent to adenine and thymine were more frequent than those adjacent to guanine and cytosine. The most prominent InDels were mononucleotide InDels. More than half of the InDels occurred within or adjacent to homopolymeric or polynucleotide repeats, and a majority of the repeats were thymine or adenine homopolymers. This study expands the current understanding of the somaclonal variation that occurs in PLB regeneration systems and long-term subcultures, which will be useful for improving the use of PLB formation in the propagation and breeding of orchids. We also confirmed that SLAF-seq is an efficient technology for genome-wide analysis of somaclonal variations for most species without reference genome sequences.
Key message
We expanded the understanding of somaclonal variation in PLB regeneration and long-term subcultures of orchids, and confirmed SLAF-seq is efficient for genome-wide analysis of somaclonal variations without reference genome sequences.
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Data availability
The Illumina SLAF-seq data developed from Oncidium are available at the NCBI Sequence Read Archive database with study accession number SRP159117.
Abbreviations
- PLB:
-
Protocorm-like body
- RFLP:
-
Restriction fragment length polymorphism
- RAPD:
-
Randomly amplified polymorphic DNA
- DNA:
-
Deoxyribonucleic acid
- AFLP:
-
Amplified fragment length polymorphism
- ISSR:
-
Inter simple sequence repeat
- SSR:
-
Simple sequence repeat
- NGS:
-
Next-generation sequencing
- SNP:
-
Single nucleotide polymorphism
- InDel:
-
Insertion-deletion
- SLAF-seq:
-
Specific-locus amplified fragment sequencing
- RRL:
-
Reduced representation library
- GC:
-
Guanine-cytosine
- Ti:
-
Transition
- Tv:
-
Transversion
- A:
-
Adenine
- T:
-
Thymine
- G:
-
Guanine
- C:
-
Cytosine
- cDNA:
-
Complementary deoxyribonucleic acid
- PCR:
-
Polymerase chain reaction
- MS:
-
Murashige–Skoog
- BA:
-
N6-benzyladenine
- NAA:
-
α-Naphthalene acetic acid
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Acknowledgements
This study was funded by the National Natural Science Foundation of China (Grant No. 31301801) and the Fundamental Research Funds for the Central Non-profit Research Institution of CAF (Grant No. CAFYBB2014QB012). The authors are grateful for the assistance provided by Biomarker Technologies (Beijing, China) in data processing.
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CXW and MT conceived the study and designed the experiments. CXW performed the experiments, analyzed the data and wrote the manuscript. MT revised the manuscript. YZ and TO helped in culturing the plant materials. All authors read and approved the final manuscript.
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Wang, C., Tian, M., Zhang, Y. et al. Molecular spectrum of somaclonal variation in PLB-regenerated Oncidium revealed by SLAF-seq. Plant Cell Tiss Organ Cult 137, 541–552 (2019). https://doi.org/10.1007/s11240-019-01589-4
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DOI: https://doi.org/10.1007/s11240-019-01589-4