MGI sequencing platforms: High-throughput gene sequencers, DNBSEQ™ sequencing technology-MGI-Leading Life Science Innovation

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	Sequencers	Sequencers	Sequencers	Sequencers	Sequencers	Sequencers	Sequencers	Sequencers
Product Model	DNBSEQ-T20×2	DNBSEQ-T10×4RS	DNBSEQ-T7	DNBSEQ-T7* For HotMPS Only	DNBSEQ-G400	DNBSEQ-G400* For HotMPS Only	DNBSEQ-G50	DNBSEQ-G99
Features	Ultra-high Throughput	Whole Process Customization	Ultra-high Daily Throughput	Ultra-high Daily Throughput	Flexible Throughput, Diverse Reads	Flexible Throughput, Diverse Reads	Flexible Data Output	Fast
Applications	Ultra-high-depth Whole Genome Sequencing	Ultra-high-depth Whole Genome Sequencing	Deep Whole Genome Sequencing	Deep Whole Genome Sequencing	WGS, WES, Transcriptome sequencing, etc.	WGS, WES, Transcriptome sequencing, etc.	Small whole genome sequencing, targeted panels, low-pass whole genome sequencing	Targeted oncology panel sequencing, infectious disease sequencing, oncology methylation sequencing
Max. Flow Cell / Run	6	8	4	4	2	2	1	2
Flow Cell Type	Slide	Slide	FC	FC	FCS & FCL	FCL	FCS & FCL	FC
Lane/Flow Cell++	1 Lane	1 Lane	1 Lane	1 Lane	4 or 2 Lanes	4 Lanes	1 Lane	1 Lane
Operation Mode	Ultra-high throughput	Ultra-high throughput	High and Ultra-high Throughput	High and Ultra-high Throughput	Medium and High Throughput	Medium and High Throughput	Medium Throughput	Low and Medium Throughput
Max. Throughput / Run	~72Tb	76.8Tb	6Tb	4Tb	1440Gb	720Gb	150Gb	48Gb
Effective Reads / Flow Cell	35B（PE100）	32B (PE150)	5000M	5000M	300M, 550M, 1500-1800M	1500-1800M	100M, 500M	80M
Average run time	60~80 hours	96~106 hours	~24 hours	20~22 hours	FCS: 13~37 hours FCL: 14~109 hours	15.5~50.5 hours	9~40 hours	5~12 hours
Min. Read Length	PE100	PE100	PE100	PE100	SE50	SE50	SE50	SE100
Max. Read Length	PE150	PE150	PE150	PE100	PE300	PE100	PE150	PE150

Recommendation of Application

Application Type	Application	Sample Type	Suitable Sequencer
Reproductive Health	NIPT	Peripheral blood	DNBSEQ-G50, DNBSEQ-G99
	PGS	Single cell	DNBSEQ-G50, DNBSEQ-G99
	CNV	Amniotic fluid, villi, abortion tissue	DNBSEQ-G50
Whole genome sequecing	WGS	Blood, saliva, gDNA	DNBSEQ-G400, DNBSEQ-G400 FAST, DNBSEQ-T7
Whole Exome Sequencing	WES	Blood, saliva, gDNA	DNBSEQ-G400, DNBSEQ-G400 FAST, DNBSEQ-T7, DNBSEQ-G99
Microbial detection	Pathogen Fast Identification	Blood, Cerebrospinal fluid, Alveolar lavage fluid	DNBSEQ-G50, DNBSEQ-G400 FAST, DNBSEQ-G99
	Gut Microbiota	Stool Specimen	DNBSEQ-G50, DNBSEQ-G400 FAST, DNBSEQ-G99
	Microbial identification and resistant analysis	Blood culture	DNBSEQ-G50, DNBSEQ-G99
Tumor	WES total solution	FFPE, Tissue, Blood	DNBSEQ-G400, DNBSEQ-G400 FAST
	Hotspot gene detection	FFPE, Tissue	DNBSEQ-G50, DNBSEQ-G400 FAST, DNBSEQ-G99
	BRCA1/2 detection	Blood, Saliva, Cervical cells	DNBSEQ-G50, DNBSEQ-G400 FAST, DNBSEQ-G99
	Lung cancer, ALK, NTRK1, NRG1, RET, ROS1, FGFR3, Fusion, Variation, etc.	FFPE, Tissue	DNBSEQ-G50, DNBSEQ-G400 FAST, DNBSEQ-G99
Plant	Molecular Breeding	Tissue, gDNA	DNBSEQ-G400, DNBSEQ-G400 FAST
Plant	Plant WGS	Tissue, gDNA	DNBSEQ-G400, DNBSEQ-G400 FAST, DNBSEQ-T7
RNA	Transcriptome Sequencing	Tissue, Cell, gDNA, RNA	DNBSEQ-G400, DNBSEQ-G400 FAST, DNBSEQ-T7, DNBSEQ-G99
RNA	RNA-Seq	Tissue, Cell, gDNA, RNA	DNBSEQ-G400, DNBSEQ-G400 FAST, DNBSEQ-T7, DNBSEQ-G99
Forensic	Individual identification	Blood, hair, Saliva, etc	DNBSEQ-G400, DNBSEQ-G400 FAST, DNBSEQ-G99
Forensic	Paternity Testing	Blood, Hair, Saliva, etc	DNBSEQ-G50, DNBSEQ-G99

DNBSEQ<span class="tm">TM</span> Technology

DNBSEQTM Technology

It features the low error accumulation of DNB and the high signal density of regular array chips to greatly improve the accuracy of detection. It has specific performance in applications such as WGS/WES with low repetition rate of output data and reduction of data waste

INTRODUCTION TO MGI SEQUENCING TECHNOLOGY

MGI's DNA sequencing instruments utilize the state-of-the-art core technology called DNBSEQ^TM. DNBs (DNA nanoballs) are pumped with by the fluidics system and loaded onto a Patterned Array chip. Sequencing primer is then added and hybridized to the adaptor region of the DNB. The sequencing reaction starts by pumping sequencing reagents containing fluorescently labeled dNTP probes and DNA polymerase. Images are taken after the fluorescently labeled probes on the DNB are excited with lasers. The images are then converted into a digital signal using MGI's propriety software. This information is then used to determine the DNA sequence of the sample.

All sequencing technologies relating to DNBs are part of DNBSEQ^TM. It includes: DNA single strand circularization and DNB preparation technology, Patterned Arrays, DNB loading, cPAS (combinatorial Probe Anchor Synthesis), Pair-End Sequencing technology on DNBs, fluidics and imaging systems, base calling algorithms, etc. cPAS technology has been widely used on various sequencing platforms including BGISEQ-50, BGISEQ-500, DNBSEQ-G50, DNBSEQ-G400, DNBSEQ-T7, etc.

Compared to other existing sequencing platforms, DNBSEQ^TM sequencing technology combines the advantages from low amplification error rates from DNBs and high density patterned arrays. These advantages dramatically improve sequencing accuracy, and have much lower duplication rates in WGS/WES applications. When combined with the PCR-free library construction method, DNBSEQ^TM also has higher SNP and indel detection accuracy compared to other platforms. In addition, the index hopping rate in DNBSEQ^TM platforms is much lower as compared to that of other platforms.

DNB Preparation Technology
Patterned Array and DNB Loading
cPAS
Pair-End Sequencing Technology
Base Call

Preparation Technology

DNB Preparation technology includes DNA single strand circularization and DNB making.

DNA single strand circularization

DNA single strand circularization: double stranded DNA with adapter sequences at the terminal ends is heated to denature and generate ssDNA (single stranded DNA). A splint oligonucleotide with a complementary sequence to both the 5’ and 3’ terminal ends of one strand of the target dsDNA will hybridize to both the 5’ and 3’ terminal ends of the same target ssDNA to form a nicked circle (Figure 1). The nick is then repaired using DNA ligase to form a single stranded circle.

DNB Making

DNA nanoballs are generated by rolling circle amplification (RCA) using the single stranded circle as a template. Various sizes of DNA fragments were amplified to roughly 100 to 1000 copies (Figure 2). DNB concentration can easily be quantified with Qubit measurements before loading onto the sequencing chip. No expensive quantification instrument or reagents are required.The primary benefit of rolling circle amplification (RCA) is the reduction in error introduced during amplification. RCA utilizes a very high-fidelity DNA polymerase, and each amplification uses the original copy of the DNA circle as the template. This makes it almost impossible to have amplification errors in the same position for all 100-1000 copies of a DNB. In addition, RCA technology avoids the exponential accumulation of errors, GC biases and dropouts observed with other amplification methods, such as PCR. All results in greatly improved sequencing accuracy with the DNBSEQ platform。

Patterned Array

Using a state-of- art semiconductor manufacturing process, a patterned binding site is created on the surface of a silicon chip. The distance between active spots on the chip surface is uniform and each binding site is only large enough to bind one single DNB. This ensures there is no interference between the fluorescence signals from neighboring DNBs. This results in high sequencing accuracy, high chip utilization, as well as optimal reagent usage.

DNB Loading

DNBs carry a negative charge in acidic conditions due to its phosphate backbone while the slide surface carries a positive charge. This positive and negative interaction is the main driving force behind DNBs loading onto the slide surface. The proprietary loading buffers can further ensure DNBs sticking on the same spot for hundreds of cycles without any compromised signals.
DNBs are optimized so they are the same size as the active sites on the slide surface. This ensures that only a single DNB is loaded onto each active site, which improves effective spot yield.

cPAS

cPAS Technology: After sequencing primers are hybridized to the adapter region of the DNB, a fluorescently labeled dNTP probe is incorporated with a DNA polymerase (Figure 4). Any unbound dNTP probes are then washed away, DNB Flow Cell is imaged (Figure 4: Imaging), fluorescence signal is converted to digital signal, and the base information is determined using MGI's proprietary base-calling software. After the image is taken, regeneration reagent is added to remove the fluorescent dye and prepares the DNBs for the next cycle

The sequencing reaction time has been reduced to less than one minute due to significant improvements in sequencing biochemistry, as well as the identification of a superior sequencing polymerase screened from tens of thousands of mutants.

2nd strand Preparation

After finishing the 1st strand sequencing, the 2nd strand generation primers and a polymerase with strand displacement activity are added to initiate 2nd strand synthesis. The polymerase will extend the new primer until it reaches the original sequenced strand, at which point it will displace the original sequencing strand to form a new single stranded template. The newly generated 2nd strand is optimized to maximize the length of the strand while ensuring the strand remains attached to the original DNB. After the 2nd strand sequencing primer is hybridized, the same sequencing chemistry is used for 2nd strand sequencing as was used for 1st strand sequencing (Figure 5). The new 2nd strand template has many more copies of insert DNA which yields much stronger signal and increased sequencing accuracy for the 2nd strand.

Base Calling Algorithm

Base calls and base call quality is calculated based on the signal intensities from all channels. The relationship between signal characterization and sequencing error is well established based on known data models. Predicted sequencing errors for unknown samples are calculated based on signal characterization. Quality scores are based on phred-33 standard.

MGI has developed a propriety Sub-pixel Registration algorithm, which enables image intensity extraction at the sub-pixel level, and greatly improves base call accuracy.

Our industry-leading technology has dramatically increased data processing speed and accuracy through integration of a GPU accelerated algorithm, optimization of execution efficiency, and real time image analysis and base calling.

Sequencers

Recommendation of Application