Guidance

Central Sequencing Laboratory: user manual 2024

Updated 1 February 2024

Introduction

The UK Health Security Agency (UKHSA) is a public health body focused on health protection and security. It prevents threats by deploying the full weight of analytic and genomic capability, on infectious diseases. It responds to threats we face with speed and scale, including terrorist threats to health, another pandemic or environmental hazards.

The Central Sequencing Laboratory (CSL) provides a centralised sequencing service of pathogens for customers across UKHSA laboratories and external to UKHSA. It provides generic nucleic acid sequencing and fragment-sizing services. The unit receives nucleic acids, plasmids and PCR products. Samples are processed based on specific protocols, and service type is selected by customers from a list in both the user manual and a memorandum of understanding (MOU) provided to each customer. Sequence and fragment-sizing data is checked by CSL against internal standards and quality scores before electronically notifying and reporting the results to allow customers to analyse their own data.

The unit has a strong research and development commitment to improve methods for rapid and high-throughput genomics. Established approaches are published and shared with microbiology specialists through training, shared protocols and workshops.

Disclaimer

This document has been controlled under the UKHSA Document Control System. Any printed copy becomes an uncontrolled document and is not managed under the UKHSA Document Control System. It is the responsibility of the copy holder to ensure that any hard copy or locally held copy in their possession reflects the current version available from the UKHSA internet site.

Amendment history

Version number	Date	Sections affected	Pages affected
10	January 2016	Change of unit name and reformatted to improve layout and access of information	All
11	April 2017	NGS run-only MiSeq service, NGS TAT, NGS repeat policy, Quality assurance section, NGS concentrations, NGS sample numbers	14 to 18
12	November 2017	Content page, NGS recharging, Sanger service codes	3, 9, 16
13	April 2019	Unit Head changed, updated link to Sanger LIMS, NGS TAT increased to 80%	6,14
14	July 2020	Unit name changed to CSL, Unit Head and deputy unit head contact details updated. Duplicated information removed throughout. Turnaround times altered for most Sanger and Fragment analysis services and NGS increased to 90% met overall. Following services removed S5B, S9, RO-1, T3, T4 and T5.	All
15	October 2021	Transfer to UKHSA template. Remove reference of Public Health England. Updated Introduction. Formatting.
16	December 2023	Email addresses updated from PHE to UKHSA. NGS sample submission web addressed changed to new SLIMS version. UKAS testing laboratory changed to UKAS medical laboratory. NGS services updated to include new run only services as well as urgent service.

Key personnel and contact details

Dr Jon Hubb
Head of CSL
[email protected]
020 8327 6068 or 07773 002652

Dr Kirstin Edwards
Deputy Head of CSL/Complaints Manager
[email protected]
020 8327 6469

Sanger sequencing/fragment analysis enquiries
[email protected]
020 8327 7772

NGS enquiries
NGS[email protected]
020 8327 7898

Sanger sequencing/fragment analysis submission

NGS submission

CSL (outside working hours)
07766 421765

UK HSA Colindale switchboard
020 8200 4400

Postal address:
The Central Sequencing Laboratory
UKHSA
61 Colindale Avenue
London
NW9 5EQ

DX address:
CSL
UK HSA Colindale
DX 6530002

CSL services

CSL offers various levels of DNA sequencing for laboratories of the UKHSA and externally (the latter by prior arrangement).

Services are provided as agreed in an MOU that outlines the terms and details of a mutual understanding or agreement. Further information can be obtained by contacting CSL.

Each sequencing request accepted by the laboratory is considered to be an agreement for work under UKHSA terms and conditions of business.

CSL offers the following sequencing services for microbial pathogens:

• Sanger sequencing of short DNA fragments on Applied Biosystems capillary platforms. Up to 850 bases read length is generated
• fragment analysis using Applied Biosystems capillary platforms to size fluorescently labelled PCR products for variable number tandem repeat (VNTR) analysis, microsatellites, ribotyping and CRISPRS (Clustered Regularly Interspaced Short Palindromic Repeat)
• next generation sequencing (NGS) using Illumina short read sequencing technology to sequence genomic and plasmid DNA or PCR amplicons

Technical specifications of services provided can be obtained in the customer MOU (described above).

Hours of service

Normal laboratory opening times are between 9am and 5pm, Monday to Friday (excluding bank holidays).

CSL provides emergency out-of-hours Sanger sequencing and fragment analysis for samples on request. A request for this service should be placed by communicating with the CSL Unit Head or Deputy Unit Head.

CSL does not offer diagnostic services to members of the public. Use of services must be agreed through prior contact and MOU agreement. There are no clinical facilities at UKHSA Colindale and we are unable to see patients or give telephone medical advice directly to members of the public.

Sample submission guidelines

DNA samples should be provided in molecular biology grade water or a TRIS-containing (EDTA free) buffer.

Submission of samples for Sanger sequencing and NGS requires the creation of a user account for online registration of samples. Prior to sample submission, customers can create accounts using the hyperlinks described in the contact details above and can register samples.

Delivery and transportation of samples

For customers within the Colindale site, deliver the samples to level 2, Zone D and place inside the refrigerator marked “CSL customer fridge”. The CSL customer sheet (BW0158), kept next to the fridge, must be completed to ensure samples are processed with the priority of when they were submitted.

For customers external to the Colindale site, samples should be sent to CSL addresses.

Arrangements must be made by referring laboratories to ensure that time and temperature requirements for sample transportation are maintained. Failure to achieve this may compromise sample integrity and the validity of test results. Samples which do not meet the sample acceptance criteria may not be processed.

Preferably samples should be sent at 2°C to 8°C or on dry ice. Samples which are dispatched at ambient temperature (10°C to 25°C) must have a transit time of no more than 72 hours. If the date of receipt is greater than 72 hours from the date of dispatch, the referring laboratory will be informed and the specimens may not be processed.

Medico-legal samples

The unit has capabilities to test medico-legal specimens. However, the unit is not accredited for performing these tests for forensics work.

Due to the legal requirements pertaining to these types of specimens, they will only be processed if the unit has been contacted in advance and if all paperwork (including the chain of evidence form) is correctly completed. This will enable the unit to ensure continuity of evidence throughout testing. The requesting laboratory must supply a Health and Care Professions Council Registered Clinical or Biomedical Scientist to monitor the CSL workflow during processing and countersign the chain of evidence form.

All requests for forensic tests must be discussed with the relevant units prior to sending the specimen to the laboratory.

Sample rejection criteria

Samples will be rejected if:

• they are not submitted in the designated 96-well plate
• their volume is different from that specified for each service
• their concentration is not within the specified range
• the online submission form has not been completed
• the plate layout does not march online submission form
• sample or primer names in the order form contain the characters or spaces \ / : * ? “ < > or the vertical bar
• a unique purchase order number has not been provided in the order submission (non-UK HSA customer orders only)

Service details

CSL offers 3 main services which are described below in Table 1.

Table 1: Sequencing services

Service code	Description of service	Turnaround time	Read length/data generated	Sample volume	DNA concentration
Sanger sequencing
S1	Reaction-ready samples (DNA and primer)	24 hours	Long (850bp)	6µl	1-100ng
S4A	PCR products for clean-up and sequencing	36 hours	Long (850bp)	10 -50µl	1-100ng
S6A	Urgent Sequencing	8 hours	Long or short (850/550bp)	6µl	1-100ng
S6B	Urgent PCR product clean-up and sequencing	10 hours	Long or short (850/550bp)	10 -50µl	1-100ng
S7A	Emergency out-of-hours sequencing	8 hours	Long or short (850/550bp)	6µl	1-100ng
S7B	Emergency out-of-hours PCR product clean-up and sequencing	10 hours	Long or short (850/550bp)	10 -50µl	1-100ng
S8A	Reaction-ready samples (DNA and primer)	24 hours	Short (550bp)	6µl	1-100ng
S8C	PCR products for clean-up and sequencing	36 hours	Short (550bp)	10 -50µl	1-100ng
CE-S1	Chain of evidence reaction ready long-read sequencing	24 hours	Long (850bp)	6µl	1-100ng
CE-S8A	Chain of evidence reaction ready short-read sequencing	24 hours	Short (550bp)	6µl	1-100ng
Fragment analysis
F4	Ready to run polymorphic size-based DNA fragment separation (for example VNTR, MLVA, ribotyping, CRISPRs)	24 hours	.fsa files for customer analysis	11.5µl	Not applicable
F5	PCR reactions for electrophoresis separation requiring pre-processing	24 hours	.fsa files for customer analysis	10µl	Not applicable
F6A	Urgent F4 service	6 hours	.fsa files for customer analysis	11.5µl	Not applicable
F6B	Urgent F5 service	7 hours	.fsa files for customer analysis	10µl	Not applicable
F7A	Out-of-hours F4 and F5 service	7 hours	.fsa files for customer analysis	10-11.5µl	Not applicable
Next-generation sequencing
N1	Standard genomic NGS	7 days	≥150Mb trimmed fastQ data	Minimum 60µl	1-100ng/µl
N2	Standard amplicon-based NGS	7 days	≥40Mb trimmed fastQ data	Minimum 60µl	1-100ng/µl
T1	Sequencing and data check	7 days	≥150Mb trimmed fastQ data	Minimum 60µl	1-100ng/µl
T3	Run only (NextSeq 1000) CSL supplied cartridge	7 days	Not applicable	Minimum 600µl	Not applicable
T4	Run only (NextSeq 550, 300 cycle) CSL supplied cartridge	7 days	Not applicable	Minimum 600µl	Not applicable
T5	Run only (NextSeq 550, 150 cycle) CSL supplied cartridge	7 days	Not applicable	Minimum 600µl	Not applicable
T6	Run-only (MiSeq) CSL supplied cartridge	7 days	Not applicable	Minimum 600µl	Not applicable
T7	Run only (NovaSeq SP) CSL supplied cartridge	7 days	Not applicable	Minimum 600µl	Not applicable
T8	Out of hours urgent NextSeq 1000 (up to 94 samples)	7 days	≥150Mb trimmed fastQ data	Minimum 60µl	1-100ng/µl

Turnaround times

Turnaround times (TAT) start from the day CSL receipts the sample in the laboratory to when data is available to the customer. Orders submitted after 4pm for Sanger sequencing or fragment analysis services and after 9.30am for NGS orders submitted may not be receipted until the following working day. The times shown are the typical TAT achieved by the laboratory but may be affected due to a surge in urgent or high-priority samples received for sequencing from an outbreak or, due to an unavoidable interruption to CSL facilities. Customers will be informed of major delays to TAT.

Specialist and technical support

CSL will provide support in designing upstream workflow to help ensure compatibility with sequencing reagents.

CSL will provide support in sequencing or fragment analysis related troubleshooting and help with interpretation of results.

Financial arrangements

For UKHSA partners please contact CSL to sign up to a MOU, which will include current pricing information. Then samples sent in the prescribed CSL format will have cost recovered via internal recharging to the partner’s project or cost centre on a monthly basis.

There will be an annual price review and this will be discussed with the partner at the same time as a review of the memorandum. All repeats will be done free of charge if the error is deemed to be due to CSL.

Please note that for non-UKHSA partners a valid Purchase Order Number must accompany each order submission. For any other queries from non-UKHSA partners, please contact CSL.

The unit will aim to give 3 months’ notice before changes are made to the price of services.

Quality assurance

An overview of quality standards at UKHSA’s laboratory services at Colindale, can be found at Quality standards: microbiology services.

CSL is a UKAS accredited medical laboratory No. 8727 in accordance with International Standard ISO 15189:2012 ‘Medical laboratories – requirements for quality and competence’. CSL will continue to maintain its accreditation and will inform its customers of any changes to accreditation status.

The schedule of accreditation listing accredited services is available on the UKAS website at Search UKAS accredited organisations (lab reference 8727).

Internal quality assurance (IQA) and external quality assurance (EQA) are carried out by partner laboratories. Results of IQA and EQA are discussed at unit and management review meetings and action taken as appropriate.

Complaints procedure

If there is a problem, or you are not satisfied with the CSL service you have received, in the first instance contact the Complaints Manager or the Unit Head. Contact details are given under Key personnel and contact details. We endeavour to be responsive to the changing needs of all users of our services.

We welcome comments on how we can improve the provision of these services. Please contact the unit if you have any queries.