PMID- 28547825
OWN - NLM
STAT- MEDLINE
DCOM- 20180402
LR  - 20210109
IS  - 2059-2310 (Electronic)
IS  - 2059-2302 (Print)
IS  - 2059-2302 (Linking)
VI  - 90
IP  - 2
DP  - 2017 Aug
TI  - Dual redundant sequencing strategy: Full-length gene characterisation of 1056 
      novel and confirmatory HLA alleles.
PG  - 79-87
LID - 10.1111/tan.13057 [doi]
AB  - The high-throughput department of DKMS Life Science Lab encounters novel human 
      leukocyte antigen (HLA) alleles on a daily basis. To characterise these alleles, 
      we have developed a system to sequence the whole gene from 5'- to 3'-UTR for the 
      HLA loci A, B, C, DQB1 and DPB1 for submission to the European Molecular Biology 
      Laboratory - European Nucleotide Archive (EMBL-ENA) and the IPD-IMGT/HLA 
      Database. Our workflow is based on a dual redundant sequencing strategy. Using 
      shotgun sequencing on an Illumina MiSeq instrument and single molecule real-time 
      (SMRT) sequencing on a PacBio RS II instrument, we are able to achieve highly 
      accurate HLA full-length consensus sequences. Remaining conflicts are resolved 
      using the R package DR2S (Dual Redundant Reference Sequencing). Given the 
      relatively high throughput of this strategy, we have developed the semi-automated 
      web service TypeLoader, to aid in the submission of sequences to the EMBL-ENA and 
      the IPD-IMGT/HLA Database. In the IPD-IMGT/HLA Database release 3.24.0 (April 
      2016; prior to the submission of the sequences described here), only 5.2% of all 
      known HLA alleles have been fully characterised together with intronic and UTR 
      sequences. So far, we have applied our strategy to characterise and submit 1056 
      HLA alleles, thereby more than doubling the number of fully characterised 
      alleles. Given the increasing application of next generation sequencing (NGS) for 
      full gene characterisation in clinical practice, extending the HLA database 
      concomitantly is highly desirable. Therefore, we propose this dual redundant 
      sequencing strategy as a workflow for submission of novel full-length alleles and 
      characterisation of sequences that are as yet incomplete. This would help to 
      mitigate the predominance of partially known alleles in the database.
CI  - (c) 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.
FAU - Albrecht, V
AU  - Albrecht V
AUID- ORCID: 0000-0001-6304-3989
AD  - DKMS Life Science Lab, Dresden, Germany.
FAU - Zweiniger, C
AU  - Zweiniger C
AD  - DKMS Life Science Lab, Dresden, Germany.
FAU - Surendranath, V
AU  - Surendranath V
AUID- ORCID: 0000-0003-1593-6461
AD  - DKMS Life Science Lab, Dresden, Germany.
FAU - Lang, K
AU  - Lang K
AD  - DKMS Life Science Lab, Dresden, Germany.
FAU - Schofl, G
AU  - Schofl G
AUID- ORCID: 0000-0003-3000-3205
AD  - DKMS Life Science Lab, Dresden, Germany.
FAU - Dahl, A
AU  - Dahl A
AD  - Deep Sequencing Group, CRTD - Center for Regenerative Therapies Dresden, Dresden, 
      Germany.
FAU - Winkler, S
AU  - Winkler S
AD  - DNA Sequencing, Max Planck Institute of Molecular Cell Biology and Genetics, 
      Dresden, Germany.
FAU - Lange, V
AU  - Lange V
AUID- ORCID: 0000-0002-6442-9573
AD  - DKMS Life Science Lab, Dresden, Germany.
FAU - Bohme, I
AU  - Bohme I
AD  - DKMS Life Science Lab, Dresden, Germany.
FAU - Schmidt, A H
AU  - Schmidt AH
AD  - DKMS Life Science Lab, Dresden, Germany.
AD  - DKMS, Tubingen, Germany.
LA  - eng
PT  - Journal Article
DEP - 20170525
PL  - England
TA  - HLA
JT  - HLA
JID - 101675570
RN  - 0 (HLA Antigens)
SB  - IM
MH  - *Alleles
MH  - *Databases, Nucleic Acid
MH  - *Genetic Loci
MH  - HLA Antigens/*genetics
MH  - High-Throughput Nucleotide Sequencing/*methods
MH  - Humans
PMC - PMC6084308
OTO - NOTNLM
OT  - HLA typing
OT  - NGS
OT  - PacBio
OT  - full-length gene sequencing
OT  - novel HLA alleles
COIS- The authors have declared no conflicting interests.
EDAT- 2017/05/27 06:00
MHDA- 2018/04/03 06:00
PMCR- 2018/08/09
CRDT- 2017/05/27 06:00
PHST- 2016/09/05 00:00 [received]
PHST- 2017/04/19 00:00 [revised]
PHST- 2017/05/01 00:00 [accepted]
PHST- 2017/05/27 06:00 [pubmed]
PHST- 2018/04/03 06:00 [medline]
PHST- 2017/05/27 06:00 [entrez]
PHST- 2018/08/09 00:00 [pmc-release]
AID - TAN13057 [pii]
AID - 10.1111/tan.13057 [doi]
PST - ppublish
SO  - HLA. 2017 Aug;90(2):79-87. doi: 10.1111/tan.13057. Epub 2017 May 25.