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Hotspot Selective Preference of the Chimeric Sequences Formed in Multiple Displacement Amplification
Jing Tu1, Na Lu2, Mengqin Duan3
1State Key Lab of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, China. jtu@seu.edu.cn.
International Journal of Molecular Sciences
|March 2, 2017
Summary
Multiple displacement amplification (MDA) creates chimeric DNA reads. Analyzing these chimeras reveals preferences in overlap length and GC content, offering strategies to minimize artifacts and distinguish them from disease-related sequences.
Area of Science:
- Genomics and Molecular Biology
- Biotechnology and Bioinformatics
Background:
- Multiple displacement amplification (MDA) is a standard method for amplifying low-input DNA.
- Chimeric reads generated during MDA can complicate genomic analyses, including structural variation and recombination studies.
- MDA by-products offer insights into the φ29 polymerase reaction mechanism.
Purpose of the Study:
- To analyze the characteristics and preferences of chimeric reads generated by MDA.
- To identify factors influencing chimera formation to optimize MDA protocols.
- To differentiate MDA-induced chimeras from disease-associated chimeric sequences.
Main Methods:
- Analysis of 36.7 million chimeric reads and 196 billion chimeric hotspots in the human genome.
- Evaluation of chimera and hotspot distribution across chromosomes.
- Assessment of preferences in overlap length, GC content, and segment distance in chimera formation.
Main Results:
- No significant chromosomal preference for chimera or hotspot distribution was observed.
- Hotspots with 12-13 nucleotide overlaps were preferentially selected for chimera generation.
- Preferences in overlap length and GC content correlate with sequence denaturation temperature, suggesting optimization targets.
- A preferred distance of 80-280 nucleotides was found between chimera segments.
Conclusions:
- Understanding MDA chimera characteristics, particularly overlap length and GC content preferences, can guide protocol optimization to reduce artifacts.
- The identified preferences provide a basis for distinguishing MDA-generated chimeras from disease-related chimeric sequences.
- This characterization enhances the reliability of MDA in various genomic applications.
