BioMicroCenter:ShortRead

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HOME -- SEQUENCING -- LIBRARY PREP -- HIGH-THROUGHPUT -- COMPUTING -- OTHER TECHNOLOGY

The MIT BioMicro Center offers short-read sequencing on five platforms from three manufacturers, spanning low to high output. All sequencing includes library quality control (RT-PCR and Fragment Analyzer), demultiplexing, and FASTQ delivery. Pricing is available to MIT users at MIT:Pricing and to all users at BioMicroCenter:Pricing.

Choosing a platform? Jump to platform comparison and FAQ below, or contact biomicro@mit.edu.

OUTPUT TIERS

OUTPUT	PLATFORM	READS / UNIT	MIN SUBMISSION	TYPICAL USE
HIGH	NovaSeqX Plus	1.2B reads/lane (10B FC) 3.0B reads/lane (25B FC)	110 µL at 2 nM per lane	Largest experiments requiring billions of reads: cohort-scale WGS, ultra-deep sequencing, high-replicate RNA-seq studies
	Available through collaboration with nearby academic shared resources. Contact BMC staff to inquire.
MID	Element AVITI24	~400M reads/lane (~800M per 2-lane flowcell)	15 µL at 2 nM	Standard depth experiments: RNA-seq, ChIP-seq, ATAC-seq, WGS at moderate coverage; tolerates low-complexity libraries well
LOW	Singular G4	~100M reads/lane (4 lanes/flowcell)	12 µL at 4 nM	Smaller experiments, amplicons, validation, pilot studies, and applications where cost-per-lane is the primary driver
	MiSeq i100	5M – 100M reads/flowcell	40 µL at 2 nM
	MiSeq classic	1M – 25M reads/flowcell	40 µL at 2 nM
	MiSeq classic is off contract and will be retired upon instrument failure. MiSeq i100 offers walkup service (MIT only, training required).

Illumina

The BioMicro Center operates the MiSeq i100 and provides access to NovaSeqX Plus through collaboration with nearby academic core facilities. Both instruments use Illumina's two-color XLEAP-SBS chemistry (G bases are inferred from signal absence) and are best suited to high-complexity libraries. UDIs are strongly recommended on both platforms.

NovaSeqX Plus

NovaSeqX Plus sequencing is available through BMC's collaboration with local academic core facilities, and is well suited to projects requiring the deepest coverage or largest sample counts. It is only available for MIT users. These collaborations provide advantaged pricing to MIT users.

• See Pricing for rates.
• Full flowcells (8 lanes, 10B or 25B) have priority over individual lanes; wait times for single lanes can vary.
• Minimum 2 nM; 110 µL per lane.
• Custom sequencing primers: at least 30 µL at 100 µM of each primer type per flowcell. Contact biomicro@mit.edu before planning a custom library prep.

MiSeq i100

Service MiSeq i100 Sequencing INPUT Illumina libraries MIN VOLUME 40 µL MIN CONCENTRATION 2 nM (~0.4 ng/µL for 300 bp library) INCLUDED SERVICES Quality Control: Fragment Analyzer and qPCR Illumina Sequencing Demultiplexing DATA FORMATS FASTQ BCL (stored 30 days) PRICING MIT:Pricing · BioMicroCenter:Pricing SUBMISSION MIT (Assisted) – iLabs MIT (Walkup) – iLabs calendar (training required)

The MiSeq i100 uses XLEAP-SBS chemistry on a patterned (CMOS nanowell) flowcell and is the BMC's walkup-capable instrument. It supports the longest read lengths of any instrument in the center (up to 1000 nt), making it ideal for amplicon panels, 16S rRNA surveys, validation runs, and other applications where read length matters more than throughput. Kits available: 100 nt (25M, 100M), 300 nt (5M, 25M, 100M), 600 nt (5M, 25M, 50M), 1000 nt (25M). Run times range from under 4 hours (25M 100 nt) to ~24 hours (100M 300 nt). Walkup service is not available to external users. Runs must be scheduled in the iLabs calendar. Server training and storage access on BMC servers is required. Custom sequencing primers: At least 15 µL of each primer type at 100 µM must be provided per flowcell. Contact biomicro@mit.edu before planning a custom library prep. Thanks to: MIT VPR, Drs. Gene Li, Michael Birnbaum, Dept. of Biology, Scott Ritterbush '89 SM '92

MiSeq classic

The MiSeq classic uses 4-color SBS chemistry on a non-patterned (random cluster) flowcell. It is available for users with established protocols on v2/v3 chemistry. We maintain minimal reagent stocks so turn around times may vary.

Kit	Approx. reads	Notes
70 nt (V2)	~15M	30 or 70 SE
150 nt (V3)	~25M	75 PE
300 nt (V2)	~15M	150 PE
500 nt (V2)	~15M	250 PE
600 nt (V3)	~25M	300 PE
500 nt NANO	~1M	Reduced output run

See MIT:Pricing · BioMicroCenter:Pricing for kit pricing. Custom sequencing primers follow the same submission guidelines as the MiSeq i100: at least 15 µL at 100 µM per primer type.

Element Biosciences

The BioMicro Center operates an AVITI24 from Element Biosciences. The AVITI24 uses Element's proprietary Avidity Base Chemistry (ABC), a 4-color sequencing approach that achieves high base-call accuracy (>80% of reads Q40+) by leveraging multivalent binding of avidites rather than single-nucleotide incorporation. The random flowcell and rolling-circle amplification approach produce low levels of index hopping and optical duplicates compared to patterned-flowcell instruments. Beyond standard NGS, the AVITI24 supports Teton CytoProfiling for direct in-cell co-detection of RNA, protein, and morphology — see Spatial Services for details.

AVITI24

Service Element Sequencing SUBMISSION Illumina (P5/P7) libraries MIN VOLUME 15 µL MIN CONCENTRATION 2 nM INCLUDED SERVICES Quality Control: Fragment Analyzer and qPCR Element Sequencing Demultiplexing ADDITIONAL SERVICES Quality Control Element library preparation DATA FORMATS FASTQ (stored 1 year) QUALITY CONTROL FASTQC Basic run metrics (alignment rate, complexity) Basic RNAseq metrics (where applicable) Basic paired end metrics (where applicable) Contamination checks PRICING MIT:Pricing · BioMicroCenter:Pricing SUBMISSION MIT – iLabs

Kits and run times: 75 PE (150 nt total): ~24 hours, ~400M reads/lane 150 PE (300 nt total): ~38 hours, ~400M reads/lane 300 PE (600 nt total): ~60 hours, ~200M reads (single lane only) Each flowcell has two independently-addressable lanes. Both lanes must be loaded to run a flowcell, but each can carry a different library pool. 300PE kits are single-lane only. Anchors: The BMC runs P5/P7-anchored (Illumina Freestyle) libraries as standard. Element Adept (SP5/SP27) libraries are also supported but require a 5′-phosphate on the forward strand for circularization — contact biomicro@mit.edu before preparing Adept libraries. Indexing: Index orientation differs from standard Illumina: indexes are read from antisense primers with the P7-side first (index 1). Provide index sequences as read on the instrument, not as ordered. For dual-indexed libraries this is typically antisense to the ordered oligo sequence. Custom primers: 70 µL at 100 µM for Read 1; 40 µL at 100 µM for all other reads. Custom primers must not interfere with other users' samples in the same lane. Minimum read guarantee: Guaranteed if (a) BMC performed QC, (b) library diversity is high especially in the first 5 bases of Read 1, and (c) concentration ≥ 2 nM. Thanks to: Element Biosciences; Scott Ritterbush

Singular Genomics

The BioMicro Center operates a G4 from Singular Genomics. The G4 uses a 4-color SBS chemistry on a patterned flowcell and delivers index-hopping rates comparable to non-patterned Illumina instruments (HiSeq/GAIIx level) — substantially lower than X-Amp patterned flowcells. This makes it a practical choice for experiments that do not require UDIs. The G4 supports four independently-addressable lanes per flowcell and up to four flowcells per run, offering per-lane flexibility at competitive per-lane cost.

G4

Service Singular G4 Sequencing INPUT Illumina (P5/P7) libraries MIN VOLUME 12 µL MIN CONCENTRATION 4 nM INCLUDED SERVICES Quality Control: Fragment Analyzer and qPCR Pool conversion: P5/P7 → S1/S2 by PCR Singular Sequencing Demultiplexing ADDITIONAL SERVICES Quality Control Singular library preparation DATA FORMATS FASTQ (stored 1 year) QUALITY CONTROL FASTQC Basic run metrics (alignment rate, complexity) Basic RNAseq metrics (where applicable) Contamination checks PRICING MIT:Pricing · BioMicroCenter:Pricing SUBMISSION MIT – iLabs

Kits and run times: 50 PE (100 nt total): ~11 hours, ~100M reads/lane 150 PE (300 nt total): ~24 hours, ~100M reads/lane Library submission: Submit standard Illumina (P5/P7) libraries. The BMC converts the pool to S1/S2 anchors by PCR prior to sequencing — do not submit S1/S2-anchored libraries directly. Indexing: After conversion, indexes are read from S1 (index 1) then S2 (index 2), in the same orientation as ordered in classical Illumina library preps. Dual 8-nt indexes are required for all lane-by-lane sequencing. Custom index primers are not permitted. Lane by lane: Must be 50 PE or 150 PE dual-indexed. Pool must be compatible with TruSeq, Nextera, smRNA, or Solexa sequencing primers. 4 lanes per flowcell; up to 4 flowcells per run. Minimum read guarantee: Guaranteed if (a) BMC performed QC, (b) no custom primers, and (c) concentration ≥ 4 nM. Thanks to: Singular Genomics

PLATFORM COMPARISON

SPEC	NovaSeqX Plus	Element AVITI24	Singular G4	MiSeq i100	MiSeq classic
Read lengths	100 nt, 300 nt	75 PE, 150 PE, 300 PE	50 PE, 150 PE	100–1000 nt	70–600 nt
Flowcell type	Patterned (X-Amp)	Random	Patterned	Patterned (CMOS nanowell)	Random
Chemistry	2-color XLEAP-SBS (G=dark)	4-color avidity SBS	4-color SBS	2-color XLEAP-SBS (G=dark)	4-color SBS
Index hopping	High — UDIs required	Low — UDIs not needed	Low (patterned, non-XLEAP)	Present — UDIs recommended	Low (non-patterned)
Low-complexity libraries	Struggles	Handles well	Handles well	Struggles	Handles well
Lane flexibility	Lane-by-lane (full FC priority)	Lane-by-lane	Lane-by-lane	Full flowcell only	Full flowcell only
Walkup	No (collaboration)	No	No	Yes (training required)	No
Illumina lib compatible	Yes (native)	Yes (P5/P7 Freestyle)	Yes — BMC converts pool by PCR	Yes (native)	Yes (native)
Donated / supported by	Local core collaboration	Element Biosciences; Scott Ritterbush	Singular Genomics	MIT VPR; Drs. Gene Li, Michael Birnbaum; Dept. of Biology; Scott Ritterbush '89 SM '92

FAQ

My library has low complexity. Which platform should I use?

Avoid the NovaSeqX Plus and MiSeq i100 — both use 2-color XLEAP-SBS chemistry where G bases are inferred from signal absence, making low-diversity early cycles prone to run failure. The AVITI24, Singular G4, and MiSeq classic all use 4-color chemistry and handle low-complexity libraries well. Always contact biomicro@mit.edu before submitting unconventional libraries.

Do I need unique dual indexes (UDIs)?

• NovaSeqX Plus – Yes, required (patterned X-Amp flowcell).
• MiSeq i100 – Recommended (patterned CMOS flowcell).
• AVITI24 – Not required; index hopping is low on random flowcells.
• Singular G4 – Not required; patterned but does not use X-Amp chemistry; hopping is on par with HiSeq/GAIIx.
• MiSeq classic – Not required; non-patterned flowcell has low hopping.

How long is my data stored?

• FASTQ files: 1 year for AVITI24, Singular G4, MiSeq i100.
• BCL files (MiSeq): 30 days.
• Long-term storage is available; see BioMicroCenter:Pricing for active storage and tape archive options.

What quality control is included?

All runs include a final pool QC (RT-PCR and Fragment Analyzer) and post-sequencing QC (FastQC, run metrics, alignment rate, complexity, RNAseq metrics where applicable, contamination checks). Users who opt out of pre-sequencing QC by providing their own concentration forfeit read guarantees.

Can I pool samples from multiple labs?

Yes, but users are responsible for minimizing index cross-contamination and index crosstalk. The BMC does not combine projects within a lane.

SUBMISSION

All sequencing is submitted through iLabs. External users should contact biomicro@mit.edu directly. Samples must meet the minimum volume and concentration requirements listed above; coordinate with BMC staff before sending non-standard submissions.