Product Information

pdfOligoMix® is a versatile, innovative, custom product for genomics discoveries. We synthesize thousands of oligonucleotide sequences in massive parallel on a microarray chip and then cleave the oligos, releasing them into solution in a single microtube. Synthesis occurs via standard DMT chemistry assuring efficient stepwise yield and a high quality final product. The product is delivered as a pool in a single microtube – ready for use in your experiment.

  1. Economical – At less than 0.8¢ per base, OligoMix is about 20 times more cost and time efficient than conventional oligos. Delivered in a single microtube, it enables inexpensive genome-scale experiments.
  2. Customizable – Customers can specify each oligonucleotide sequence (lengths up to 150-mers). We can synthesize oligonucleotides in OligoMix containing labels, such as terminus phosphate, amino and thiol with linkers, biotin, FAM or other dyes.
  3. Reliable – Innovative microfluidic array platform ensures high quality synthesis. Multiple QC steps are implemented at various stages of OligoMix manufacturing. OligoMix is subjected to both hybridization and qRT-PCR assays to assess final quality.
  4. Simple & Fast – Download our excel spreadsheet order form, paste in your sequences and email back to us. Product can be delivered in 1-2 weeks.

Microfluidic Array Platform—in situ Synthesis

OligoMix® achieves high synthesis purity because it is produced via an advanced microarray synthesis technology (µParaflo®) that integrates a photo-generated acid (PGA) chemistry, digital photolithography (DLP), and advanced microfluidics to enable high throughput parallel synthesis of custom DNA microarrays. The PGA chemistry enables the use of standard oligo building blocks, and eliminates the need for any specially modified nucleotides which may exhibit lower coupling efficiency. DLP technology enables programmable synthesis of custom sequences and the µParaflo® microfluidic device contains the synthesis reactions each within a picoliter-scale reaction chamber, producing more uniform synthesis than reactions performed on the open surface of a slide.

Biomarker

  • Conventional Chemicals
  • Established Synthesis Processes

  • Efficient Stepwise Yield
  • Quality Final Product

Synthesis Technology References

  • Gao X, Yu PY, LeProust E, Sonigo L, Pellois JP, Zhang H. (1998) Oligonucleotide synthesis using solution photogenerated acids. Journal of the American Chemical Society 120, 12698-12699 [abstract].
  • Srivannavit O, Gulari M, Gulari E, LeProust E, Pellois JP, Gao X, Zhou X. (2004) Design and fabrication of microwell array chips for a solution-based, photogenerated acid-catalyzed parallel oligonucleotide DNA synthesis. Sensors and Actuators A. 116, 150-160 [abstract].
  • Zhou X, Cai S, Hong A, Yu P, Sheng N, Srivannavit O, Yong Q, Muranjan S, Rouillard JM, Xia Y, Zhang X, Xiang Q, Ganesh R, Zhu Q, Makejko A, Gulari E, Gao X. (2004) Microfluidic picoarray synthesis of oligodeoxynucleotides and simultaneously assembling of multiple DNA sequences. Nucleic Acids Research 32, 5409-5417 [abstract].
  • Tian J, Gong H, Sheng N, Zhou X, Gulari E, Gao X, Church G. (2004) Accurate multiplex gene synthesis from programmable DNA chips. Nature 432, 1050-1054 [abstract].

Targeted Sequencing

Though next-generation DNA sequencing (NGS) provides very high levels of coverage, even on complex genomes, it is still advantageous to reduce the complexity of samples and sequence smaller targeted regions – in particular, when sample numbers are very high and the goal is detection of less prevalent mutations. Several academic and commercial groups have developed a variety of capture methods for enriching or selectively amplifying subsets of the genome for targeted sequencing.

The key performance parameters of these methods are capture specificity and sensitivity, the ability to multiplex many samples and capture large regions of interest, and of course, cost. Recently developed methods include: Solution Hybrid Selection (SHS)2,9, Molecular Inversion Probes (MIP)2,10, Selective Genomic Circularization (SGC)6,11,12 and Oligo-Selective Sequencing (OS-Seq)1. All of these methods have been demonstrated to be effective at selectively enriching desired regions of interest within a given genome.

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While each method has its own advantages and disadvantages that make them suitable for specific situations5, the common thread amongst these methods is the need for high quality synthesis of large numbers of oligonucleotide sequences for use as capture probes or primers. This can prove expensive when using conventional solid support column based synthesis methods as the number of sequences can reach into the tens of thousands.

  • It has been demonstrated that the use of microarray synthesized oligos produces the required numbers and quality of oligos quite effectively at a far lower cost12.
  • OligoMix® has been demonstrated as an effective method of oligo synthesis for targeted sequencing in MIP2, SGC6, and OS-Seq1 targeted sequencing methods.

Targeted Methylation Analysis

As with most genomic analysis methods, CpG methylation analysis must be: quantitative, high-throughput, cost-effective, and both scalable and flexible with respect to coverage. Ideally, one would be able to efficiently investigate the methylation of large numbers of CpGs in large numbers of samples.

The standard method for measuring methylation involves treatment of DNA with sodium bisulfite which causes conversion of unmethylated cytosines (C) to uracils (U), whereas 5-methylcytosine (5mC)s remain unchanged. The differences in reactivity of Cs and 5mCs to bisulfite can be distinguished by subsequent microarray5 or sequencing3,4 methods.

Both of these methods can benefit from prior targeted capture and amplification of suspected CpG regions in order to reduce the complexity of samples and focus the analysis on specific genomic segments. The use of oligonucleotides for targeted capture increases both sample throughput and coverage, while decreasing cost per sample. Using an OligoMix® synthesis strategy vs. individual oligo synthesis further increases flexibility, scalability and cost efficiency of targeted methylation analysis methods. Recently, two new capture methods have been developed for targeted methylation analysis.

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One challenge for these methodologies lies in the construction of capture probe panels. They must be customizable to different genomic targets, scalable to a very large sample size (1,000–100,000 samples), and inexpensive. The current procedures are labor intensive and costly, making it impractical for construction of very large panels or custom panels. As a parallel oligo synthesis technology capable of producing virtually unlimited numbers of oligos of lengths up to 100 nucleotides as a pool, OligoMix® overcomes this barrier and represents a significantly more cost effective method for construction of probe panels than single-plex PCR.

  • Probes generated by OligoMix® were compared to single-plex PCR constructed probes using ROC analysis and no significant difference in performance was observed5.
  • Oligo pools therefore represent an inexpensive means for constructing large and custom dU probe panels and greatly improve the flexibility of the assay with respect to coverage.

 

Product Descriptionmix of DNA oligonucleotide sequences
Number of Oligosthousands of sequences or more per tube
Oligo Formsingle stranded (ss); desalted and ready for reaction
Lengthup to 150 mers (inquire for longer oligos)
5’ or 3’ Terminus Modificationsphosphate, fluorescent dyes, biotin, linkers, and others
Internal Modificationsmodified DNA or RNA bases
Yield*tens of attomoles per sequence and a total of sub-fmols per OligoMix® tube
Price(see www.lcsciences.com/oligomix)
Delivery14 days

 

References
  1. Myllykangas S, Buenrostro JD, Natsoulis G, Bell JM, Ji HP. (2011) Efficient targeted resequencing of human germline and cancer genomes by oligonucleotide selective sequencing. Nat Biotechnol 29, 1024–27.
  2. Teer JK, Bonnycastle LL, Chines PS, Hansen NF, Aoyama N, Swift AJ, Abaan HO, Albert TJ, Margulies EH, Green ED, Collins FS, Mullikin JC, Biesecker LG. (2010) Systematic comparison of three genomic enrichment methods for massively parallel DNA sequencing. Genome Res 20(10), 1420-31.
  3. Diep D, Plongthongkum N, Gore A, Fung H, Shoemaker R, Zhang K. (2012) Library-free methylation sequencing with bisulfite padlock probes. Nature Methods 9(3), 270-2.
  4. Labrie V, Buske OJ, Oh E, Jeremian R, Ptak C, Gasiūnas G, Maleckas A, Petereit R, Žvirbliene A, Adamonis K et al.(2016) Lactase nonpersistence is directed by DNA-variation-dependent epigenetic aging. Nature Structural & Molecular Biology 23(6):566-73.
  5. Nautiyal S, Carlton VE, Lu Y, Ireland JS, Flaucher D, Moorhead M, Gray JW, Spellman P, Mindrinos M, Berg P, Faham M. (2010) High-throughput method for analyzing methylation of CpGs in targeted genomic regions. Proc Natl Acad Sci 107(28), 12587-92.
  6. LC Sciences’ customers – data unpublished
  7. LC Sciences’ internal development work – data unpublished
  8. Gnirke A, Melnikov A, Maguire J, Rogov P, LeProust EM, et al. (2009) Solution hybrid selection with ultra-long oligonucleotides for massively parallel targeted sequencing. Nat Biotechnol 27, 182–89.
  9. Porreca GJ, Zhang K, Li JB, Xie B, Austin D, Vassallo SL, LeProust EM, Peck BJ, Emig CJ, Dahl F, Gao Y, Church GM, Shendure J. (2007) Multiplex amplification of large sets of human exons. Nat Methods 4(11), 931-36.
  10. Dahl F, Stenberg J, Fredriksson S, Welch K, Zhang M, et al. (2007) Multigene amplification and massively parallel sequencing for cancer mutation discovery. Proc Natl Acad Sci USA 104, 9387–92.
  11. Natsoulis G, Bell JM, Xu H, Buenrostro JD, Ordonez H, et al. (2011) A Flexible Approach for Highly Multiplexed Candidate Gene Targeted Resequencing. PLoS ONE 6(6), e21088.
  12. Baker M. (2011) Microarrays, megasynthesis. Nat Methods 8(6), 457-60.

 


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