Daniel:Notebook/RNAFISH/2014-7-7
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Probe Design
Designing probes for Matt and Hosuk's rolony project. The three genes they are testing are ACTB, RAB7A, and MALAT1. I will design probe sets for these three genes.
Workflow
- Obtain transcript sequence from Genome Browser
- Use genome browser to find genes, and tables to download transcript (mRNA only)
- Use gene sequence as input for probe designer
- Designer created by Arjun Raj, utilized by Long Cai for paper
- Creates 20mer probes
- Copy/paste probe sequences into .txt file
- Print only sequence to the oligo file
- awk '{print $2}' malat1_probes.txt > oligos_malat1.txt
- Attach primer sequences before/after
- VI command, forward: :%s!^!GTCATATCGGTCACTGTT!
- VI command, reverse: :g/$/norm AGATCAGGATACACACTACCC
- Primer sequences are AP1 and AP2 from the V6 set, used in High Resolution Chromosome Painting
Price Comparison IDT and Stellaris
IDT Primers | IDT Fluorophores | Stellaris | |
Length | 58 | 20 | 20 |
Number of Oligos | 93 | 93 | 93 |
Price/Base (100 nmol, USD) | 0.28 | 0.28 | NA |
Modifications | None | Fluorophore | Fluorophore |
Oligo Price | 1510.32 | 520.8 | 1725 |
Modifications | 0 | 85 | 0 |
Total Price | 1510.32 | 44268 | 1725 |
It would appear Stellaris can do this pretty cheap.