Matt:LabNotes/2014-6-4
Jump to navigation
Jump to search
Agi26k0gap Probes for use in Mouse Brain Tissue?[edit]
- Hosuk was wondering if we could use the Agi26k0gap probeset in the mouse brain tissue sections he has generated rolonies in
- This would allow us to test second rolony generation in fixed tissue without using human brain tissue
Procedure[edit]
- Download Mus Musculus mRNA sequences from Biomart
- Filter: Genes - protein coding only
- Build bowtie2 index
bowtie2-build Mus_proteincoding_mart_export.fa Mus_proteincoding
- Turn Dec2012_0bp_gap_probes.txt and Dec2012_0bp_gap_probes_stemCellGenes.txt into a fasta file with target sequences (concatenate columns 3 and 5)
- Align target sequences to mouse mRNA
bowtie2 -x Mus_proteincoding -f Dec2012_targetseq.fa > Targetseq_alignto_MusRNA.sam & 13127 (100.00%) were unpaired; of these: 12038 (91.70%) aligned 0 times 643 (4.90%) aligned exactly 1 time 446 (3.40%) aligned >1 times 8.30% overall alignment rate
Conclusion[edit]
- Only 8% of padlock probes would hybridize (and this doesn't even account for small number of mismatches) so Agi26k probes should not be used in mouse tissue