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如何使用PSI-BLAST:How to use the PSI-BLAST?  

2012-05-18 11:06:04|  分类: 生物信息分析 |  标签: |举报 |字号 订阅

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Position-Specific Iterated (PSI)-BLAST,是一种更加高灵敏的Blastp程序,对于发现远亲物种的相似蛋白或某个蛋白家族的新成员非常有效。当你使用标准的Blastp 比对失败时,或比对的结果仅仅是一些假基因或推测的基因序列时("hypothetical protein" or "similar to..."),你可以选择PSI-BLAST重新试试。

可以使用一下链接:PSI-BLAST

psiblast page First use the Query menu and select PSI-BLAST on any page to access the PSI-BLAST form, then follow these steps:

1. Type the "sw:UBXN1_HUMAN" protein identifier in the input text area

2. Check the Swiss-Prot database

3. Select matrix "BLOSUM45"

4. Click on the search button
psiblast result page The top part summarizes your input data and options. Then you will see small messages about the evolution of the search.
match map When the search is finished, you get the results in 3 parts.

a) the statistics (useful to observe the evolution of the collected proteins)

b) the graphic (described here)

   1. Score per residue, with highly conserved ones shown in one-letter code

   2. Position and significance of the matches

   3. Description of the matches

c) the alignments (see below)
psiblast pairwise alignment c) the alignments

   1. Detailed description of the match (position, species, scores, alignment) with links to the original entry and to a graphical view

   2. In a smaller font, a member of the same cluster (identity level above threshold)

   3. In grey non-matching alignment
send results to Check only the matches you want to keep (e.g., those matching the first 60 amino acids) and then use the drop down menu to send the matches either:

- ...next cycle (redo a PSI-BLAST cycle with new PSSM from selected matches)
- ...to PSI-BLAST form
- ...to MSA Hub (please choose this option)
How to use the MSA Hub?
msa_hub page Here you can observe your multiple sequence alignment, modify it, and send it to another page.

For example you want to use the "Catalogue".

1. Select "HTML walker [10 prot./page]" (10 proteins per page)

2. Select "shrink to fit line width"

3. Click on "Catalogue" button
catalogue result page Now you can see one image for each matching protein with the description at the top:

1. Link to original protein entry

2. Shaded box displaying the position of the match

3. Same but for another protein

4. Other features found in the protein entry

Now please click on the "back to Hub" button (top of the page), then in the MSA Hub click on the "jalview2" button
How to use Jalview?
Jalview start page Please click again on the "Start Jalview" button to launch the Java applet in another window
Jalview MSA editor/viewer This is the view of the multiple alignment given by the Jalview applet.

You can immediately notice that the matching region is only located in the first 60 amino acids. Using Jalview, please reduce the size of the alignment. (see Jalview documentation)
MSA in Jalview This is the reduced multiple sequence alignment. Once you are satisfied with it, please select the Jalview file sub-menu "Output to Browser"
What next?
Back to MSA_hub 1. The modified multiple sequence alignment is now back in the text area

2. Select the "...to PSI-BLAST" to send your alignment to the PSI-BLAST form

Comment: this alignment is very small and doesn't contain many gaps, thus you don't need to re-align the sequences. In the case of a longer and more complex alignment, we recommend using MAFFT or T-Coffee to re-align the sequences before going back to PSI-BLAST.
psiblast submission 1. The modified multiple sequence alignment is now in the text area

2. Check other databases. (Note that the BLOSUM45 is magically kept from your first search)

3. Click on the search button to submit your multiple alignment to PSI-BLAST
psiblast matches

Congratulations you have defined a protein domain, and now you get more matche
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