when I send these miRNA to mRNA database using blastn I get these result. what I would to do fined a new miRNA taht similar to the query sequence.
I run it by this command line
blastn -db esthuman.fasta -word_size 7 -query allprecursor.txt -out nasr -perc_identity 100 -outfmt 6 -max_target_seqs 2 hsa-let-7b gi|262205597|ref|NR_029479.1| 100.00 83 0 0 1 83 1 83 3e-35 154 hsa-let-7b gi|4826511|emb|AL049853.1| 100.00 83 0 0 1 83 17069 16987 3e-35 154 hsa-let-7c gi|262205602|ref|NR_029480.1| 100.00 84 0 0 1 84 1 84 8e-36 156 hsa-let-7c gi|7768689|dbj|AP001667.1| 100.00 84 0 0 1 84 186545 186628 8e-36 156 hsa-let-7d gi|262205605|ref|NR_029481.1| 100.00 87 0 0 1 87 1 87 2e-37 161 hsa-let-7d gi|71516309|gb|CH471089.1| 100.00 87 0 0 1 87 25762708 25762794 2e-37 161 hsa-let-7f-1 gi|262205612|ref|NR_029483.1| 100.00 87 0 0 1 87 1 87 2e-37 161 hsa-let-7f-1 gi|71516309|gb|CH471089.1| 100.00 87 0 0 1 87 25760221 25760307 2e-37 161 hsa-let-7g gi|262205206|ref|NR_029660.1| 100.00 84 0 0 1 84 1 84 8e-36 156 hsa-let-7g gi|71518807|gb|CH471055.1| 100.00 84 0 0 1 84 52281689 52281606 8e-36 156
please anyone who have use blast help
Dear nasromer2191989, Hi
1- why you have used "precursors" ? As it is shown in Figure 4 of the paper which @Parsad has kindly offered below, they used Mature mirna first.
2- did you "remove redundant miRNA" first from your database? (it is also shown in the fig 4)
I use the precursor because the mature it dose not showing any hits or score. do you know how to turn the subject sequence to miRNA? Thanks
by default when you run blastn it performs megablast(word size=28) this might be a reason you may not find any hits as miRNA ranges from (16-30 max). while doing blastn enable the
-task blastn-shortor else change the word size. All options can be found here
I could not run blastn for mature miRNA which command line I have to use for short sequence
could you share your blast command
Dear Parsad This is the command line I have used blastn -db mrna.fasta -query matureall.txt -out resultmature.out
And I get this result it si no hits found
Database: mrna.fasta 508,597 sequences; 15,096,209,361 total letters
No hits found
Lambda K H 1.33 0.621 1.12
Gapped Lambda K H 1.28 0.460 0.850
Effective search space used: 45261163845
No hits found
Try this version.
please Can I have your email for more discuss in blast work
I have use your suggestion for mature sequence and I did not get which sequence I have to use in Mfold. While the subject sequence is too short for folding. for instance this is the first sequence I get please can you advice me which one I have to take to MFold.
emb|LQ069844.1| Sequence 168 from Patent EP2964234 42.1 0.009
Score = 42.1 bits (21), Expect = 0.009 Identities = 21/21 (100%), Gaps = 0/21 (0%) Strand=Plus/Plus
What data source have you used for mature miRNA? Example ID is
unassigned RNAfrom human, same sequence is identified as hsa-let-7a-3p in miRBase.
For the mfold part, usually miRNAs positioning on genome is identified and upstream & downstream sequences (~50 to 100bps) from the position (including miRNA) are considered to see the stem-loop structure. You could use tools like mirdeep2, mireap etc
The data source is miRbase. I have made file of miRNAs in fasta format I want to use it as query to EST & GSS (human). By this work I need to identify miRNA similar to the one in the file. So that I compare their target.What I see in this paper (Identification of miRNA encoded by Jatropha curcas from EST and GSS ) they send the known miRNA to EST&GSS and identify new miRNA and their target.
I am confusing 1)-which sequence they use in Mfold. 2)-Are they use 100% Identities or use the one with specific number of mismatch
kindly refer methodology section of the paper you have mentioned for all the parameters.
If you are working on human data, you may use human genome instead of gss or est. see mirdeep or mireap, pretty east to implement for novel miRNA prediction
I use the human miRNA against the EST & GSS human also. I have got the mismatched less than 2 so could I use them as similar miRNA.
Please can I have your email address for more guidelines ?
Please do not ask for personal email addresses. They are not shared publicly on Biostars. We encourage users to keep all discussions open.
i do agree with @genomax2, as the discussion here would help many others
Thanks for replying
So, Could we follow the steps together? I have done blastn then I confuse which sequence I have to use for blastx.
If you have done blast of human miRNA against human EST. Filter miRNA-EST pairs with the cutoff mentioned in the paper. Then predict secondary structure of entire EST. if proper secondary structure forms (stem-loop) take those ESTs for blastx to remove protein coding. details steps and parameters are mentioned in the paper
please how did you find the name of this miRNA (hsa-lat-7a-3p) by the information in the result file.
I took embl id from here, extracted sequence and did blast in miRBase. Are trying to do similar study as in paper or you have sequenced miRNA data?
I have miRNA data I collected from miRbase. I need to find the similar to what I have then I check their target. please advise me If there is tool can preform it. The tool allow me to scan my file to any other data base and give me back the similar miRNA.
as i had mentioned earlier you could refer mirdeep2 (pipeline tool). If you want web tools, miRNAkey, miRanalyzer, sRNAtoolbox
Here you can find a list of other tools 1, 2
How did you blast (emb|LQ069844.1| Sequence 168 from Patent EP2964234 Length=21 )to miRbase I want to check the mismatched one if it is already there in miRbase.
opt search by sequence from mirbase
I want to do blastx on all the sequence that I get in result file. I want to check if is it the same miRNA. that I did blastn.