Hybridization capture is integrated into the overall next generation sequencing workflow immediately before sequencing on an NGS platform, such as Illumina. A fully sequenceable, barcoded/indexed NGS library (or pool of multiple libraries) is denatured, and allowed to anneal to complementary target-specific biotinylated probes/baits. These bait:library complexes are then bound to streptavidin-coated magnetic beads via the biotin on the probes, which are washed to remove non-specifically bound molecules. The remaining “enriched” library molecules are then released from the baits and amplified before sequencing.

Note! You may know the “hybridization capture” technique by another name, such as:

  • Target enrichment
  • Target capture
  • Probe capture
  • Exon capture
  • Capture sequencing / sequence capture
  • Hybridization sequencing / hyb-seq
  • Hybridization capture / hyb-cap

Specific recommendations for per-library input mass for different enrichment project types can be found in the applicable myBaits manual.

Target capture necessarily requires subjecting your libraries to a bottleneck, wherein target molecules are captured and therefore enriched, and non-target molecules are therefore removed. To have sufficient unique molecules for good sequencing coverage of your targets, successful captures DEPEND on the input of sufficiently complex libraries.
For best results, it is recommended that only amplified (non-PCR-free) NGS libraries are used for target capture. This provides multiple copies of each starting template molecule, increasing the chance of each individual molecule getting enriched. However if you need more starting material to reach the recommended amount, it is generally preferable to generate more library from fresh genomic DNA or a new batch of indexed library, rather than through extra amplification. This is because while some amplification is good, over-amplification risks reducing the observable complexity of your libraries through the uneven action of PCR bias, as some molecules will become relatively more abundant while others become rare. This is also true for manipulating your libraries after capture: amplify your post-capture libraries the minimum number of cycles necessary to reach the molarity required by your sequencing facility.

The applicable myBaits manual covers some common technical questions and troubleshooting topics at the end of each protocol. Please read through the relevant section first as it may answer your question. If you still have an issue, please contact us via email at techsupport_at_arbor.daicel.com or reach out to your most recent contact person for assistance.

When ordering your myBaits kit, please indicate the sequencing library configuration you intend to enrich. The standard adapter blocking reagent provided with the kit (Block X) is compatible with Illumina® TruSeq®-style or Nextera®-style libraries with single 6-12 bp or dual 6-12 bp indexing. These options cover the vast majority of currently available commercial library preparation systems intended for sequencing on any Illumina platform.

For different adapter configurations than those described above, we recommend ordering Custom IDT® xGen® Blocking Oligos. At a concentration of 1 μg/μL, custom adapter-blocking oligos can be used in lieu of myBaits Block X.
If you are not certain, or later decide to change your library prep kit, please contact us so we can instruct you on how to obtain the correct blocking oligos.

Yes, please contact sales@arbor.daicel.com to add our subsampling service to your project.

We strongly discourage sending frozen plant samples because freeze-thaw cycles are terrible for DNA integrity. Also, we have found that lyophilization has large positive effects on yield and purity for plants. If you do submit frozen material, ensure that once it is frozen, it is not allowed to thaw.

We tested a wide variety of plastics and seals before making this recommendation. This is the combination that we have identified as capable of standing up to homogenization without leaking, so we do require these specific plastics to be used. Having trouble sourcing them? Contact genomics@arbor.daicel.com for help.

Yes. If they are submitted in our required plastics, you can add our lyophilization service to your project. If you would like us to format your samples correctly for lyophilization, we can do so for an additional fee.

Yes! Check out Appendix 2 of our Sample Preparation Guide. It has tips on how to get your plant subsamples into a plate. Tried our tips and still need help? You can add our subsampling service to your project.

No, due to the potential effects of upstream handling (which we cannot control), we cannot guarantee the output mass, purity, or integrity of extracted nucleic acids. Please see our NGS Services Policies.