Processing
Alt-R HDR Donor Oligos
Built for homology-directed repair (HDR)
Our proprietary synthesis process delivers HDR-ready oligos of the highest quality, up to 200 bases. For longer HDR donors, you can order our Megamers. Alt-R HDR Donor Oligos are available in tube or plate formats.
- Ideal for introducing point mutations or short insertions
- Industry-leading HDR rates
- Design-optimized through extensive wet-bench testing and validation
- Available for human, mouse, rat, zebrafish, and C. elegans
The CRISPR basics handbook
Everything you need to know about CRISPR, from A to Z, for beginners as well as advanced users.
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HDR Positive Controls
Use the HDR positive controls below along with the Alt-R HDR Donor Oligos to determine the HDR efficiency in your experiments. HDR positive controls provide different crRNA and scale options, and include tracrRNA along with human or mouse donor oligos. See the product sheet for additional information and protocol details about the HDR positive controls in the Resources tab below.
| Guide RNA | Human (Homo sapiens) | Mouse (Mus musculus) |
|---|---|---|
| Alt-R™ CRISPR-Cas9 crRNA | Add to Cart | Add to Cart |
| Alt-R™ CRISPR-Cas9 crRNA XT | Add to Cart | Add to Cart |
| Alt-R™ CRISPR-Cas9 sgRNA | Add to Cart | Add to Cart |
Alt-R HDR Design Tool
If you don’t have a template design of your own, use our Alt-R HDR Design Tool to design your template. Simply provide basic information about your target site, and then use the tool to design and visualize your desired edit within the sequence. The Alt-R HDR Design Tool will provide the recommended HDR donor template along with gRNA(s) for your specifications.
Product details
Homology-directed repair (HDR) donors
IDT provides different HDR donor options for researchers to select from, depending on the specific needs of their application:
- No modification—standard DNA
- PS modification—4 phosphorothioate bonds: 5′–N*N*NNN…NNN*N*N–3′, where * represents a PS bond and N are DNA bases
- Alt-R HDR modification—IDT’s proprietary modification pattern developed through extensive testing to increase donor oligo stability and enable the highest rate of repair
Performance
Alt-R HDR Donor Oligos improve HDR efficiency
The HDR rates from experiments using 3 different donor options were compared. Donor oligos with Alt-R HDR modifications increase HDR rates dramatically compared to other formats and gave the highest HDR rates in the industry (Figure 1).
Figure 1. Alt-R HDR donors improve HDR efficiency over other donor types. HeLa and Jurkat cells were electroporated with 2 µM Cas9 RNP complexes (Alt-R S.p. HiFi Cas9 Nuclease V3 complexed with Alt-R CRISPR-Cas9 crRNA and tracrRNA) targeting 4 genomic loci along with 0.5 µM single-stranded HDR donor template using the Nucleofection™ System (Lonza). Donor templates contained no modifications (Unmodified), 4 phosphorothioate linkages (2 at each end of the template; PS-modified), or the Alt-R HDR modification (Alt-R HDR–modified). Genomic DNA was isolated 48 hours (HeLa) or 72 hours (Jurkat) after electroporation, and HDR efficiency was measured by amplicon sequencing on the Illumina MiSeq® system (v2 chemistry, 150 bp paired-end reads).
Alt-R HDR modified donors and Alt-R HDR Enhancer lead to the highest HDR rates
We investigated whether combining Alt-R HDR Enhancer with Alt-R modifications of HDR donor oligos would improve HDR rates further. Our data demonstrate that this works well, leading to the highest HDR rates (Figure 2).
Figure 2. Combined use of Alt-R HDR modified donors and Alt-R HDR Enhancer have an additive effect on HDR improvement. RNP complex (2 μM) targeting 3 genomic loci along with 0.5 μM single-stranded HDR donor oligo were delivered to HeLa cells by electroporation using the 4D-Nucleofector™ System (Lonza). The RNP complex comprised Alt-R S.p. HiFi Cas9 Nuclease V3 complexed with Alt-R CRISPR-Cas9 crRNA and tracrRNA. Unmodified, PS-modified, or Alt-R HDR modified donor templates were used. Immediately after electroporation, cells were plated in media with or without 30 μM Alt-R HDR Enhancer. Genomic DNA was isolated 48 hours after electroporation, and HDR efficiency was measured by amplicon sequencing on an Illumina™ MiSeq™ system (v2 chemistry, 150-bp paired-end reads).
