Genome and metagenome sequencing: Using the human methyl-binding domain to partition genomic DNA derived from plant tissues.
OBJECTIVE
Variation in the distribution of methylated CpG (methyl-CpG) in genomic DNA (gDNA) across the tree of life is biologically interesting and useful in genomic studies. We illustrate the use of human methyl-CpG-binding domain (MBD2) to fractionate angiosperm DNA into eukaryotic nuclear (methyl-CpG-rich) vs. organellar and prokaryotic (methyl-CpG-poor) elements for genomic and metagenomic sequencing projects. •


METHODS
MBD2 has been used to enrich prokaryotic DNA in animal systems. Using gDNA from five model angiosperm species, we apply a similar approach to identify whether MBD2 can fractionate plant gDNA into methyl-CpG-depleted vs. enriched methyl-CpG elements. For each sample, three gDNA libraries were sequenced: (1) untreated gDNA, (2) a methyl-CpG-depleted fraction, and (3) a methyl-CpG-enriched fraction. •


RESULTS
Relative to untreated gDNA, the methyl-depleted libraries showed a 3.2-11.2-fold and 3.4-11.3-fold increase in chloroplast DNA (cpDNA) and mitochondrial DNA (mtDNA), respectively. Methyl-enriched fractions showed a 1.8-31.3-fold and 1.3-29.0-fold decrease in cpDNA and mtDNA, respectively. •


CONCLUSIONS
The application of MBD2 enabled fractionation of plant gDNA. The effectiveness was particularly striking for monocot gDNA (Poaceae). When sufficiently effective on a sample, this approach can increase the cost efficiency of sequencing plant genomes as well as prokaryotes living in or on plant tissues.
New England Biolabs, 240 County Road, Ipswich, Massachusetts 01938 USA.
Erbay
E
Yigit
Department of Biology, New Mexico State University, P.O. Box 30001 Msc 3AF, Las Cruces, New Mexico 88003 USA.
David I
DI
Hernandez
Department of Biology, New Mexico State University, P.O. Box 30001 Msc 3AF, Las Cruces, New Mexico 88003 USA.
Joshua T
JT
Trujillo
New England Biolabs, 240 County Road, Ipswich, Massachusetts 01938 USA.
Eileen
E
Dimalanta
Department of Biology, New Mexico State University, P.O. Box 30001 Msc 3AF, Las Cruces, New Mexico 88003 USA.
C Donovan
CD
Bailey
endophyte; enrichment; genome sequencing; metagenome; methyl-binding domain; microbiome