forward: 2.5 days cluster restoring: 1 day reverse: 2.5 days
throughput (for mappable with repeats)
* may be even higher
** for MP-run
SOLiD 2 system / Applied Biosystems
optimal beads density*
mappable with repeats
mappable w/o repeats
throughput (for mappable with repeats)
* one deposition well per slide, ~300x106 beads for download
** for fragment run, MP-run should increase throughput on 10-20%
454 (FLX Titanium Serie) / Roche
throughput (filter-passed bases)
0.4-0.6Gb per run ~1Gb per day
99.5% at 250 bases 99% at 400 bases
price per raw base
~100x higher than for Illumina/SOLiD ~the same as for capillary sequencing
bead-based libraries, no UV-lamps or lasers in sequencer
According to throughput it is a next-generation sequencing system, but according to price — not. A lot of applications are not practical for 454 because of high price per nucleotide.
Comparison of Illumina and SOLiD systems
Which system is better? It is impossible to answer question in this form. At the moment both systems have their own advantages and disadvantages. To make a decision it is necessary to think about main applications of the system.
More or less clear situations:
high throughput resequencing of large genomes: SOLiD is more accurate;
RNA structure analysis: short-insertion MP-sequencing is available only for Illumina.
Let suppose, that it is necessary to resequence human genome with 10x coverage with mate-paired procedure. It is about 10 x 3Gb = 30Gb of mappable nucleotides.
time is practically the same;
more hand work for SOLiD (but one technical assistant is enough in both cases);
price (w/o equipment) is practically the same (flowcells are more expensive for Illumina, sequencing — for SOLiD);
equipment price per project is also quite similar, because run time is a bit shorter for Illumina (26 days // 30 days) and price of instrument is a bit less for SOLiD;
short-insertion MP-sequencing is impossible on SOLiD platform;
read length of long-insertion MP-sequencing is restricted to 25bases on SOLiD platform (because of the program);
error rate is much less for SOLiD. It is important for low-coverage projects. But, coverage variance of the SOLiD is about two times higher than coverage variance for Illumina, which is not good for low-coverage analysis. Besides, high variance of coverage on SOLiD makes analysis of structural variations more difficult. Taken together, combination "accuracy + coverage" is better for SOLiD, but not dramatically.
It is also possible to increase the accuracy of Illumina by eliminating reads with low quality. In this case more runs would be necessary to perform, so the overall price would increase.
Throughput: Illumina ≈ SOLiD
throughput (mappable nucleotides per day) is about the same for both systems: ~1.2 Gb/day. The equivalence is regularly disturbed after upgrading of one of the systems, but upgrading plans for both platforms are quite similar in the nearest future.
Read length (RL): Illumina ≈ SOLiD
about the same now: ~35b as a kit, ~50b as a home-made modification of protocol. Near-future plans are quite similar.
increasing of RL have both positive and negative consequences. Optimal RL is different for different applications.
Quality: Illumina < SOLiD
Accuracy of SOLiD platform is higher, because each nucleotide is analysed twice in ligation-based sequencing. As a result single-nucleotide substitution change colour of two dinucleotides. Standing alone colour changes may be filtered out as sequencing errors.
development plans for near future are quite similar for both platforms: increase of seq-elements density, better chemistry/enzymes, acceleration of imaging, etc.
long-term development is also similar. From technical point of view, both machines are combination of liquid handling robot with fluorescent scanner. If it would happen, that some particular sequencing approach is much better, than competitors, both robots might be adapted for this procedure.
if compare polymerisation and ligation approaches, it is unclear now which is more perspective, but unlikely, that perspectives are the same. In both cases progress will be limited (unlikely, that any of the systems would reach read length and quality of the capillary sequencing). But, basic principles are completely different, so one approach will reach internal limitations early, than other.
Fragment libraries preparation: Illumina ≈ SOLiD
fragment libraries for both systems are DNA fragments with adaptors attached to both ends. Any procedure/kit developed for one system may be relatively easy adapted for the other.
the same story for any single-read libraries: fragment genomic libraries, barcodes, microRNA libraries, etc.
minimal amount of starting material is about the same for both systems. Minor difference: ePCR on beads is more restrictive to the fragment length than bridge amplification. If starting material is a set of heterogenious short DNA fragments (chromatin immunoprecipitation) more fragments will be suitable for Illumina sequencing, than for the SOLiD one.
MP-libraries <600bp: Illumina only
Illumina have developed a technology for changing orientation of clusters directly on the glass. It gives a possibility to sequence library fragments from both ends. The only restriction is the length of DNA fragment. Everything is fine if fragment length is below ~300bp. Fragments in the range 300-600bp also may be sequenced, but with lower efficiency: cluster sizes are heterogeneous, so cluster density should be lower.
in case of "large-size MP-libraries" read length is limited by the length of restriction fragments. Sequencing of "small-size MP-libraries" may be done with any desirable read length.
SOLiD have no instrument for the small-size MP-sequencing:
MP-libraries can't be constructed from relatively short (<500bp) fragments, because they can't bend to form a circle;
in the very beginning Applied Biosystems claimed, that the ligation-based technology would be able to sequence fragments in both directions. But now it works only in 3'→5' direction;
problem would not solve completely, even if Applied Biosystems would develop 5'→3' sequencing or template inversion, because "beads / emulsion PCR" does not work well for fragments >200bp
MP-libraries >600bp: Illumina ≈ SOLiD
large-size MP-libraries differ only by sequence of adaptors. Any protocol may be adapted for both systems. For example, MP-library construction protocol for Illumina presented on this site is based on SOLiD procedure.
Flowcell preparation: Illumina >> SOLiD
for Illumina is about 3 times less
much more for SOLiD
small-scale library prep
Illumina: no restrictions SOLiD: it is impossible to prepare PCR emulsion for less than 1/2 of the slide (because of geometry of ULTRA TURRAX DT-20 tube)
loading of several samples
Illumina: 8 independent samples may be loaded without any loss of throughput SOLiD: 4 independent samples — loss of 1/4 of glass area; 8 samples — loss of 1/3 of area
much better for Illumina procedure
days for Illumina, months for SOLiD
Sequencing: Illumina > SOLiD
Illumina: ready-to-load kit SOLiD: about 4 hours
about the same hand-work
Illumina: one reagent loading for one-direction sequencing SOLiD: enzyme and primer plates should be changed each day; reagents should be refilled several times during the run
In house development (libraries): Illumina > SOLiD
the only restriction for the library development for Illumina: distal parts of adapters should correspond to the primers immobilized on the flowcell. Design of alternative sequencing primers is easy and straightforward.
a lot of inconveniences with SOLiD. As for Illumina, one of the adaptors should correspong to the primer immobilized on beads. It is impossible to change sequencing primer without "opening" the kit, because a special "blocking" and special "bridge" primers should be used (company neither supply information about structure of this primers, nor takes orders for custom synthesis). Besides, at least 5 different primers should be ordered for one sequencing place.
In house development (flowcell): Illumina < SOLiD
SOLiD flowcell is a plain glass. It is possible to customize bead deposition, etc. Practically nothing is possible to do inside of Illumina's channels.
In house development (sequencing): Illumina < SOLiD
modification of the instrument programm
Illumina: easy and clear both for image collection and liquid handling SOLiD: impossible*
fluorescence excitation and detection
Illumina: uses two lasers, excitation wavelength are practically fixed SOLiD: UV lamp, in principle it is possible to change filters, but it is unclare how to program
* EcoP15I MP-libraries have 27b-long fragments, but it is possible to select only (20, 25, 30, 35, 40,...) cycles in SOLiD instrument control software. Changing of read length from 25 to 27 would be really helpful. We tried to push company to modify the program, but without success. On Illumina it would take ~5min to change the number of cycles.
Kits and consumables (convenience): Illumina > SOLiD
Illumina kits: some reagents should be dispensed by 2µl pipette, but delivered in 1.7ml tubes: Klenow, Klenow exo(-), primers. Practically no other problems;
a lot of inconveniences with SOLiD kits:
misleading names of reagents:
* two completely different reagents have similar names: "Bead Wash Buffer" and "Bind & Wash Buffer";
* "Low Salt Binding Buffer" instead of "Binding Buffer";
cups without labels:
* it is difficult to handle 11-26 tubes with primers without cup-labels;
* it is possible to change cups by mistake;
labels on some tubes completely hide the content of the tube;
some reagents, that should be dispensed by 2µl pipette are delivered in large tubes;
a lot of reagents may be premixed in advance;
there are only oligonucleotide sets for library preparation, complete kits are not available;
quantities of viscous solutions (glycerol, ePCR components) should be expressed in terms of weight (not volume);
company did not check, if primer plates might be stored frozen;
Additional equipment: Illumina ≈ SOLiD
both systems require:
additional computing facilities for data analysis and storage;
room with climate control (NB, SOLiD produce more heat);
nebulizer is a bad instrument for DNA shearing: some ultrasonic device should be ordered;
Hydroshear is obligatory for preparation of large-insertion libraries;
should be enough number of 96-well PCR machines for ePCR;
Ordering and delivery: Illumina << SOLiD
Illumina: delivery takes at least 3 weeks (Europe);
SOLiD: normally 1-3 days from order to delivery. Excellent.
Stability of the system: Illumina > SOLiD
both systems are not completely stable. Sometimes (one per ~20) run fails for unknown reasons. Illumina is a little more stable than SOLiD in this respect.
Illumina uses lasers, SOLiD — UV lamp. Lasers are much more expensive, but UV-lamp has a much shorter operation life (about 0.5 year). Each UV lamp costs ~$600.
Service and support: Illumina < SOLiD
people from Applied Biosystems are ready to help in installation and training of technical assistants;
reparation is fast, broken part delivered in a next day;
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