Blast+ 使用補充筆記

摘要： Blast比對軟體大概是是短序列區域性比對軟體中最常用的一個了，但是其引數眾多，一些引數一直沒好好仔細研究過，如下： 增加blast比對結果資訊 blast的-outfmt 引數，使blastp -help 即可檢視每個輸出格式的資訊，如下所示： *** Fo...

Blast比對軟體大概是是短序列區域性比對軟體中最常用的一個了，但是其引數眾多，一些引數一直沒好好仔細研究過，如下：

增加blast比對結果資訊

blast的-outfmt 引數，使blastp -help 即可檢視每個輸出格式的資訊，如下所示：

*** Formatting options
 -outfmt <String>
alignment view options:
0 = Pairwise,
1 = Query-anchored showing identities,
2 = Query-anchored no identities,
3 = Flat query-anchored showing identities,
4 = Flat query-anchored no identities,
5 = BLAST XML,
6 = Tabular,
7 = Tabular with comment lines,
8 = Seqalign (Text ASN.1),
9 = Seqalign (Binary ASN.1),
10 = Comma-separated values,
11 = BLAST archive (ASN.1),
12 = Seqalign (JSON),
13 = Multiple-file BLAST JSON,
14 = Multiple-file BLAST XML2,
15 = Single-file BLAST JSON,
16 = Single-file BLAST XML2,
18 = Organism Report

Options 6, 7 and 10 can be additionally configured to produce
a custom format specified by space delimited format specifiers.
The supported format specifiers are:
qseqid means Query Seq-id
qgi means Query GI
qacc means Query accesion
qaccver means Query accesion.version
qlen means Query sequence length
sseqid means Subject Seq-id
sallseqid means All subject Seq-id(s), separated by a ';'
sgi means Subject GI
sallgi means All subject GIs
sacc means Subject accession
saccver means Subject accession.version
sallacc means All subject accessions
slen means Subject sequence length
qstart means Start of alignment in query
qend means End of alignment in query
sstart means Start of alignment in subject
send means End of alignment in subject
qseq means Aligned part of query sequence
sseq means Aligned part of subject sequence
evalue means Expect value
bitscore means Bit score
score means Raw score
length means Alignment length
pident means Percentage of identical matches
nident means Number of identical matches
mismatch means Number of mismatches
positive means Number of positive-scoring matches
gapopen means Number of gap openings
gaps means Total number of gaps
ppos means Percentage of positive-scoring matches
frames means Query and subject frames separated by a '/'
qframe means Query frame
sframe means Subject frame
btop means Blast traceback operations (BTOP)
staxid means Subject Taxonomy ID
ssciname means Subject Scientific Name
scomname means Subject Common Name
sblastname means Subject Blast Name
sskingdom means Subject Super Kingdom
staxids means unique Subject Taxonomy ID(s), separated by a ';'
(in numerical order)
sscinames means unique Subject Scientific Name(s), separated by a ';'
scomnames means unique Subject Common Name(s), separated by a ';'
sblastnames means unique Subject Blast Name(s), separated by a ';'
(in alphabetical order)
sskingdoms means unique Subject Super Kingdom(s), separated by a ';'
(in alphabetical order) 
stitle means Subject Title
salltitles means All Subject Title(s), separated by a '<>'
sstrand means Subject Strand
qcovs means Query Coverage Per Subject
qcovhsp means Query Coverage Per HSP
qcovus means Query Coverage Per Unique Subject (blastn only)

其實我們一般常用的就是-outfmt 5 或者-outfmt 6 ，前者輸出XML格式，後者輸出TAB分割格式；前者在早期一篇博文Blast+ xml格式解讀中提起過（資訊比較全，用處也相對比較廣），而後者則是平時最為常用的格式（也是一些軟體喜歡呼叫的格式）

TAB格式每列資訊如下（可以對照上面的說明理解一下）：

qseqid sseqid pident length mismatch gapopen qstart qend sstart send evalue bitscore

但我們有時想要的並不止上述12列資訊，比如我還想知道比對結果的覆蓋度資訊（qcovs：Query Coverage Per Subject）

其實只要在blast比對命令中先事先加上需要增加的列ID即可，如在outfmt 6 基礎上加上覆蓋度資訊：

-outfmt "6 qseqid sseqid pident length mismatch gapopen qstart qend sstart send evalue bitscore qcovs"

注：需要幾列就一直往上加即可，空格分割

分割NR子庫

之前分割NR子庫選用的是早期一篇博文ofollow,noindex" target="_blank">建立NR子庫以及從NR庫提取特定物種分類的序列

但是現在NCBI出了blast-2.8版本，其可支援用NCBI自帶程式碼分割的NR子庫的索引作為比對的庫，使用比較方便

• Support for a new version of the BLAST database that allows you to limit search by taxonomy as well some other improvements.

當然如果用這個版本的話，NR庫也要重新下載了ftp://ftp.ncbi.nlm.nih.gov/blast/db/v5/

使用方式也比較簡單（至少比之前的方法方便了），如果只想比對單一物種（如人:9606的話），命令如下：

blastp –db nr –query query.fasta –taxids 9606 –outfmt 6 –out blast.outfm6

如果想比對NR子庫哺乳動物的話，需要先建個哺乳動物子庫索引

get_species_taxids.sh -t 40674 > 40674.txids

然後再將序列比對至NR哺乳動物子庫

blastp –db nr –query query.fasta –taxidlist 40674.txids –outfmt 6 –out blast.outfm6

具體說明可看：https://ftp.ncbi.nlm.nih.gov/blast/db/v5/blastdbv5.pdf

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