Transcript process redesign in the Exeter laboratory

From kit car to Toyota production
line; process redesign in the
Exeter laboratory
Sian Ellard, Carolyn Tysoe, Martina Owens, Melissa
Sloman, Kevin Colclough, Andrew Parrish, Neil
Goodman, Karen Stals, Michael Day, Katie Guegan,
Ann-Marie Patch and Beverley Shields
Laboratory Automation
Barcoding
DNA
Extractor
Liquid handling
robots
TaqMan
LIMS
Sequencer
Process redesign
Pre White Paper
Kit car
Post White Paper
Toyota production line
Toyota Production System:
Principles of lean thinking
 Specify value
 Create flow
 Reduce waste (muda)
 Monitor performance
PCR set up
Pre White paper
Batching approach
4 or 8 patients per gene
Post White paper
Production line
Any number of patients, any gene
PCR set up
(a) Large genes/medium throughput
eg. BRCA1/2
(b) “Normal traffic”
Need to use common PCR mix and
thermocycler program
PCR workflow
Kit car
7 thermocycler programs
298 amplicons
5 PCR mixes
Toyota
MegaMix Royal
60°C anneal
426/432 amplicons
How many water blanks?
Kit car
One water blank per amplicon
Toyota
?
 Water blank contamination for manual PCRs assessed by gel
electrophoresis 1/685 (0.15%)
 Water blank contamination rate for PCRs (manual and robotic)
assessed by gel electrophoresis and sequencing; 0/1647 (<0.07%)
96-well sequencing pipeline
Manual PCR
Sequencing
by scientists and
technologists
by one technologist
ExoSAP/DyeEx
3730
Unidirectional
Exon ± 10bp
Mutation Surveyor + visual inspection
DNA extraction process
DNA Extraction
 Gentra installed March 2007
 DNA stored in 2D barcoded
tubes since March 2007
DNA
Quantification
Samples in 2D
bar coded
tubes
 Software for barcode checking
of samples on/off Gentra
October 2008; Excel installed
January 2009 and system
implemented in February
Robotic PCR workflow
DNA in 2D
barcoded tubes
Primer dilutions in
2D barcoded tubes
Check rack layout
Check rack layout
MegaMix
Normalised DNA plate (96 well)
PCR plates (4x96 well)
384-well sequencing pipeline
Robotic PCR
Sequencing
Band 4 technologists
Band 4 technologist
Agencourt
3730
Unidirectional
Exon ~-50 to +10bp
Mutation Surveyor (semi-automated)
Semi-automated sequence analysis
 Sensitivity of unidirectional analysis for heterozygous
substitutions is >99.6% (n=701 heterozygous base
substitutions) CMGS study; Genetic Testing and Molecular Biomarkers In press
 Inspect mutation report to confirm mutations and
polymorphisms (delete false positive calls)
 Inspect HGVS table to check ROIs covered, ROI quality
scores meet threshold and inspect bases with a PHRED-like
score <20
Mutation report
HGVS (Quality) report
Rationalisation of workflows
 Triplet repeats – rationalised within SCOBEC
 LightCycler assays (FVL, prothrombin and HFE) to TaqMan. Robotic
setup and result generated from software.
 Agarose gel assays (B27 and JAK2) replaced by fluorescent PCR (added
to HNF1A c.872dup assay). Robotic setup and result generated from
GeneMarker software.
 Manual tests: Haemato-oncology, CFTR OLA and MLPA
Aim of increased laboratory
automation
 Safer systems
 Increased efficiency
 Increased capacity
Activity data 2001- 2008
14000
12000
Number
10000
Samples
8000
Reports
6000
Genotypes
4000
2000
0
01-02
02-03
03-04
04-05
05-06
06-07
07-08
92% of genotypes are now generated by sequencing (vs 36% in 2001-2002)
Reporting time data 2005 - 2009
2500
100%
90%
2000
80%
70%
1500
60%
50%
1000
40%
30%
500
20%
10%
0
0%
2005-2006
2006-2007
2007-2008
2008-2009
Projected
40 days (no. of reports)
40 days (% within target)
Implementation of laboratory
automation
Barcoding
DNA
Extractor
Liquid handling
robots
TaqMan
Sequencer
SCOBEC Genetics Laboratories Network
Molecular genetic testing
Past
Present
Future