The Center currently hosts a G4 platform by Singular Genomics. Singular’s G4 platform is a short-read mid-throughput sequencing platform delivering adequate sequencing data through its proprietary sequencing-by-synthesis chemistry at a significantly competitive price point.
Singular requires S1/S2 anchors in place of Illumina P5/P7 anchors. These can be replaced 1:1 in most oligo designs:
s1:ACAAAGGCAGCCACGCACTCCTTCCCTGT
s2:CTCCAGCGAGATGACCCTCACCAACCACT
INDEXING
Singular indexes are read from the Anchor sequences in all cases and custom primers for indexes are not allowed.
Indexes are read from the S1 primer first (index1) followed by S2 (index2)
Index reads are the SAME SEQUENCE as ordered in most classical library preps.
Indexes are REQUIRED and should be 8 nucleotides in all lane by lane sequencing.
Min custom primer submission: 20 uL at 100 uM
LANE BY LANE
Must be 50 or 150 paired-end dual indexed
Pool must be compatible with TruSeq, Nextera, TruSeq, smRNA and Solexa sequencing primers
MINIMUM READS
the BMC has performed quality control.
No custom primers.
submitted libraries are at least 2 nM.
All other requests, including use of custom sequencing primers require a full flowcell. Illumina libraries which have been converted in the Center will also require a full flowcell.
The G4 Platform
SPEC
G4
SEQUENCER
READS/LANE Low number is minimum per lane for standard libraries
F3 per Lane: 50-100 M
RUN TIME
50 PE 11 hours
150 PE 24 hours
KITS AVAILABLE
100nt
300nt
KEY NOTES
4-color chemistry
Patterned flow cell
4-lanes per flow cell
4-flow cells per run
Anchor replacement or anchor expansion of Illumina libraries
THANKS TO
Singular Genomics
INDEX HOPING
ACCURACY
Index hopping for a standard set of Singular libraries
Percent Perfect Plot for Singular libraries 150PE
Index hopping on Singular runs is on par with GAIIx and HiSeq. High levels of index hopping as observed on Illumina X-AMP chemistry is not observed.
Percent perfect plot for phiX on current Singular chemistry (Manley et al., 2016)
