PMID- 29494651
OWN - NLM
STAT- MEDLINE
DCOM- 20180622
LR  - 20190221
IS  - 1932-6203 (Electronic)
IS  - 1932-6203 (Linking)
VI  - 13
IP  - 3
DP  - 2018
TI  - A streamlined workflow for single-cells genome-wide copy-number profiling by 
      low-pass sequencing of LM-PCR whole-genome amplification products.
PG  - e0193689
LID - 10.1371/journal.pone.0193689 [doi]
LID - e0193689
AB  - Chromosomal instability and associated chromosomal aberrations are hallmarks of 
      cancer and play a critical role in disease progression and development of 
      resistance to drugs. Single-cell genome analysis has gained interest in latest 
      years as a source of biomarkers for targeted-therapy selection and drug 
      resistance, and several methods have been developed to amplify the genomic DNA 
      and to produce libraries suitable for Whole Genome Sequencing (WGS). However, 
      most protocols require several enzymatic and cleanup steps, thus increasing the 
      complexity and length of protocols, while robustness and speed are key factors 
      for clinical applications. To tackle this issue, we developed a single-tube, 
      single-step, streamlined protocol, exploiting ligation mediated PCR (LM-PCR) 
      Whole Genome Amplification (WGA) method, for low-pass genome sequencing with the 
      Ion Torrent platform and copy number alterations (CNAs) calling from single 
      cells. The method was evaluated on single cells isolated from 6 aberrant cell 
      lines of the NCI-H series. In addition, to demonstrate the feasibility of the 
      workflow on clinical samples, we analyzed single circulating tumor cells (CTCs) 
      and white blood cells (WBCs) isolated from the blood of patients affected by 
      prostate cancer or lung adenocarcinoma. The results obtained show that the 
      developed workflow generates data accurately representing whole genome absolute 
      copy number profiles of single cell and allows alterations calling at resolutions 
      down to 100 Kbp with as few as 200,000 reads. The presented data demonstrate the 
      feasibility of the Ampli1 WGA-based low-pass workflow for detection of CNAs in 
      single tumor cells which would be of particular interest for genome-driven 
      targeted therapy selection and for monitoring of disease progression.
FAU - Ferrarini, Alberto
AU  - Ferrarini A
AD  - Menarini Silicon Biosystems spa, Bologna, Italy.
FAU - Forcato, Claudio
AU  - Forcato C
AD  - Menarini Silicon Biosystems spa, Bologna, Italy.
FAU - Buson, Genny
AU  - Buson G
AD  - Menarini Silicon Biosystems spa, Bologna, Italy.
FAU - Tononi, Paola
AU  - Tononi P
AD  - Menarini Silicon Biosystems spa, Bologna, Italy.
FAU - Del Monaco, Valentina
AU  - Del Monaco V
AD  - Menarini Silicon Biosystems spa, Bologna, Italy.
FAU - Terracciano, Mario
AU  - Terracciano M
AD  - Menarini Silicon Biosystems spa, Bologna, Italy.
FAU - Bolognesi, Chiara
AU  - Bolognesi C
AD  - Menarini Silicon Biosystems spa, Bologna, Italy.
FAU - Fontana, Francesca
AU  - Fontana F
AD  - Menarini Silicon Biosystems spa, Bologna, Italy.
FAU - Medoro, Gianni
AU  - Medoro G
AD  - Menarini Silicon Biosystems spa, Bologna, Italy.
FAU - Neves, Rui
AU  - Neves R
AD  - Department of General, Visceral and Pediatric Surgery, Medical Faculty, 
      University Hospital of the Heinrich- Heine-University Dusseldorf, Dusseldorf, 
      Germany.
FAU - Mohlendick, Birte
AU  - Mohlendick B
AD  - Department of General, Visceral and Pediatric Surgery, Medical Faculty, 
      University Hospital of the Heinrich- Heine-University Dusseldorf, Dusseldorf, 
      Germany.
FAU - Rihawi, Karim
AU  - Rihawi K
AD  - Unita Operativa di Oncologia Medica, Policlinico Sant'Orsola - Malpighi, Bologna, 
      Italy.
FAU - Ardizzoni, Andrea
AU  - Ardizzoni A
AD  - Unita Operativa di Oncologia Medica, Policlinico Sant'Orsola - Malpighi, Bologna, 
      Italy.
FAU - Sumanasuriya, Semini
AU  - Sumanasuriya S
AD  - The Institute of Cancer Research and Royal Marsden NHS Foundation Trust, London, 
      United Kingdom.
FAU - Flohr, Penny
AU  - Flohr P
AD  - The Institute of Cancer Research and Royal Marsden NHS Foundation Trust, London, 
      United Kingdom.
FAU - Lambros, Maryou
AU  - Lambros M
AD  - The Institute of Cancer Research and Royal Marsden NHS Foundation Trust, London, 
      United Kingdom.
FAU - de Bono, Johann
AU  - de Bono J
AD  - The Institute of Cancer Research and Royal Marsden NHS Foundation Trust, London, 
      United Kingdom.
FAU - Stoecklein, Nikolas H
AU  - Stoecklein NH
AD  - Department of General, Visceral and Pediatric Surgery, Medical Faculty, 
      University Hospital of the Heinrich- Heine-University Dusseldorf, Dusseldorf, 
      Germany.
FAU - Manaresi, Nicolo
AU  - Manaresi N
AD  - Menarini Silicon Biosystems spa, Bologna, Italy.
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20180301
PL  - United States
TA  - PLoS One
JT  - PloS one
JID - 101285081
SB  - IM
MH  - Adenocarcinoma/genetics
MH  - Adenocarcinoma of Lung
MH  - Cell Line, Tumor
MH  - DNA Copy Number Variations
MH  - Female
MH  - High-Throughput Nucleotide Sequencing/instrumentation/*methods
MH  - Humans
MH  - Lung Neoplasms/genetics
MH  - Male
MH  - Neoplasms/*genetics
MH  - Neoplastic Cells, Circulating/pathology
MH  - Polymerase Chain Reaction/instrumentation
MH  - Prostatic Neoplasms/genetics
MH  - Single-Cell Analysis/instrumentation/*methods
MH  - Whole Genome Sequencing/instrumentation/*methods
MH  - Workflow
PMC - PMC5832318
COIS- Competing Interests: AF, CF, GB, CB, VDM, PT, MT, FB, FF, GM, NM are employee of 
      Menarini Silicon Biosystems S.p.A.. GB, PT, NM are co-inventors on the patent 
      "Method and kit for the generation of dna libraries for massively parallel 
      sequencing" (WO 2017178655 A1) on the method presented, assigned to Menarini 
      Silicon Biosystems S.p.A. The method presented has been implemented as a kit 
      called *Ampli*1 LowPass, commercially available from Menarini Silicon Biosystems 
      S.p.A.. This does not alter our adherence to PLOS ONE policies on sharing data 
      and materials.
EDAT- 2018/03/02 06:00
MHDA- 2018/06/23 06:00
PMCR- 2018/03/01
CRDT- 2018/03/02 06:00
PHST- 2017/11/23 00:00 [received]
PHST- 2018/02/19 00:00 [accepted]
PHST- 2018/03/02 06:00 [entrez]
PHST- 2018/03/02 06:00 [pubmed]
PHST- 2018/06/23 06:00 [medline]
PHST- 2018/03/01 00:00 [pmc-release]
AID - PONE-D-17-41362 [pii]
AID - 10.1371/journal.pone.0193689 [doi]
PST - epublish
SO  - PLoS One. 2018 Mar 1;13(3):e0193689. doi: 10.1371/journal.pone.0193689. 
      eCollection 2018.