Artificial polyploidy improves bacterial single cell genome recovery.
BACKGROUND
Single cell genomics (SCG) is a combination of methods whose goal is to decipher the complete genomic sequence from a single cell and has been applied mostly to organisms with smaller genomes, such as bacteria and archaea. Prior single cell studies showed that a significant portion of a genome could be obtained. However, breakages of genomic DNA and amplification bias have made it very challenging to acquire a complete genome with single cells. We investigated an artificial method to induce polyploidy in Bacillus subtilis ATCC 6633 by blocking cell division and have shown that we can significantly improve the performance of genomic sequencing from a single cell.


RESULTS
We inhibited the bacterial cytoskeleton protein FtsZ in B.subtilis with an FtsZ-inhibiting compound, PC190723, resulting in larger undivided single cells with multiple copies of its genome. qPCR assays of these larger, sorted cells showed higher DNA content, have less amplification bias, and greater genomic recovery than untreated cells.


CONCLUSIONS
The method presented here shows the potential to obtain a nearly complete genome sequence from a single bacterial cell. With millions of uncultured bacterial species in nature, this method holds tremendous promise to provide insight into the genomic novelty of yet-to-be discovered species, and given the temporary effects of artificial polyploidy coupled with the ability to sort and distinguish differences in cell size and genomic DNA content, may allow recovery of specific organisms in addition to their genomes.
Department of Energy Joint Genome Institute, Los Alamos National Laboratory, Los Alamos, New Mexico, United States of America.
Armand E K
AE
Dichosa
Michael S
MS
Fitzsimons
Chien-Chi
CC
Lo
Lea L
LL
Weston
Lara G
LG
Preteska
Jeremy P
JP
Snook
Xiaojing
X
Zhang
Wei
W
Gu
Kim
K
McMurry
Lance D
LD
Green
Patrick S
PS
Chain
J Chris
JC
Detter
Cliff S
CS
Han
D001412:Bacillus subtilis / Q000166:cytology* / Q000187:drug effects / Q000235:genetics*; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D004269:DNA, Bacterial / Q000378:metabolism; D016680:Genome, Bacterial / Q000235:genetics*; D023281:Genomics / Q000379:methods*; D011123:Polyploidy; D011725:Pyridines / Q000494:pharmacology; D059010:Single-Cell Analysis / Q000379:methods*; D013844:Thiazoles / Q000494:pharmacology
D004269:DNA, Bacterial; C531944:PC190723; D011725:Pyridines; D013844:Thiazoles