FastQ格式介绍

为了便于测序数据的发布和共享，高通量测序数据以FASTQ 格式来记录所测的碱基读段和质量分数．如下图 所示，FASTQ 格式以测序读段为单位存储，每条读段占4 行，其中第1 行和第3行由文件识别标志和读段名(ID)组成(第1 行以“@”开头而第3 行以“+”开头；第3 行中ID 可以省略，但“+”不能省略)，第2 行为碱基序列，第4行为对应的测序质量分数．

FastQ数据格式

1.序列名称：

对于每一条FastQ序列，都有一个可以唯一标示的序列名称，如下：
 [code lang="text"]
 @HWUSI-EAS100R:6:73:941:1973#0/1
 [/code]
HWUSI-EAS100R
the unique instrument name
6
flowcell lane
73
tile number within the flowcell lane
941
'x'-coordinate of the cluster within the tile
1973
'y'-coordinate of the cluster within the tile
#0
index number for a multiplexed sample (0 for no indexing)
/1
the member of a pair, /1 or /2 (paired-end or mate-pair reads only)  
 
Versions of the Illumina pipeline since 1.4 appear to use #NNNNNN instead of #0 for the multiplex ID, where NNNNNN is the sequence of the multiplex tag.
With Casava 1.8 the format of the '@' line has changed:
[code lang="text"]
 @EAS139:136:FC706VJ:2:2104:15343:197393 1:Y:18:ATCACG
 [/code]
EAS139
the unique instrument name
136
the run id
FC706VJ
the flowcell id
2
flowcell lane
2104
tile number within the flowcell lane
15343
'x'-coordinate of the cluster within the tile
197393
'y'-coordinate of the cluster within the tile
1
the member of a pair, 1 or 2 (paired-end or mate-pair reads only)
Y
Y if the read fails filter (read is bad), N otherwise
18
0 when none of the control bits are on, otherwise it is an even number
ATCACG
index sequence
2、质量值：对于每一条序列，其每一个碱基都有一个对应的测序质量值：
传统测序的质量值是基于Phred quality scores，定义如下：
Phred quality scores Q are defined as a property which is logarithmically related to the base-calling error probabilities P.
Q=-10 log10P
 Phred quality scores are logarithmically linked to error probabilities
Phred Quality Score
Probability of incorrect base call
Base call accuracy
10
1 in 10
90 %
20
1 in 100
99 %
30
1 in 1000
99.9 %
40
1 in 10000
99.99 %
50
1 in 100000
99.999 %
The Solexa pipeline (i.e., the software delivered with the Illumina Genome Analyzer) earlier used a different mapping, encoding the odds p/(1-p) instead of the probability p:
Although both mappings are asymptotically identical at higher quality values, they differ at lower quality levels (i.e., approximately p > 0.05, or equivalently, Q < 13).
为了便于序列存储，通常采用单字符来标示序列的质量值。至于序列的quality values值，是通过一些算法得出来的。即：用字母的ASCII值减去相应的数（不同测序平台数值不一样），然后就得到Q值，然后通过前面的计算公式计算出碱基的测序错误率。
下面是不同测序平台使用的字符区间段：