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===BLAST RT RAB7A primer sequences against rRNA=== *From [[Matt:LabNotes/2014-1-31#RT_RAB7A Primers and the Number of RAB7A Primary Rolonies]] it seems like some sequences are complementary to some really high abundance transcripts ====BLAST with ribosomal RNA==== *Used Biomart sequence retrieval tool, searched for unspliced (transcript) and used Filter = Gene: rRNA with Associated Gene Name as heading for fasta file **Downloaded and stored in my genome-miner as RNAsequences.fa in /RT_RAB7A *Built database formatdb -i RNAsequences.fa -p F *Saved a fasta file of RT primer sequences and then BLAST-ed with wordsize of 7, 8, 9, and 10 blastall -i RT_RAB7A_primers.txt -d RNAsequences.fa -p blastn -o result_wordsize7.txt -W 7 blastall -i RT_RAB7A_primers.txt -d RNAsequences.fa -p blastn -o result_wordsize8.txt -W 8 ... *Shortened primer sequences to 7 by removing from the 5'end and BLAST-ed with wordsize of 7 blastall -i primers_7nt.txt -d RNAsequences.fa -p blastn -o result_7.txt -W 7 ====Results==== *BLAST of 10nt primers **Wordsize 7: [[File:Result_wordsize7.txt]] **Wordsize 8: [[File:Result_wordsize8.txt]] **Wordsize 9: [[File:Result_wordsize9.txt]] **Wordsize 10: [[File:Result_wordsize10.txt]] *BLAST of 7nt primers: [[File:Result_7.txt]] ====Conclusion==== *Looking only at primers that match at the 3' end, the only alignments are to RNA pseudogenes **1000nt RT primer has NO alignments of any kind *Therefore it is unlikely that it is because of high amounts of rRNA primary rolonies that 1000nt shows so much signal
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