Bcl to fastq conversion problem in bcl2fastq-2.17

Question

[BioRyder]

Hello All,

I am facing some some problem while trying to convert bcl to fastq by using bcl2fastq-2.17 software.Partial generation of output data is the problem. R1 and R2 reads are generated for lane 4.But for Lane1,2,3, only Read2 generated and no data for other lanes.While checking the Report folder there are status report present for each lane.It will be a great help for me if any body can help me to find out the problem. Hiseq2500 is used for generating the data.

The following command I used for bcl to fastq conversion.

nohup bcl2fastq \
  --runfolder-dir /data/coredata/genomics/hiseq/150827_SN1046_0229_BC6WPEACXX/ \
  --output-dir /data/coredata/genomics/hiseq/150827_SN1046_0229_BC6WPEACXX/TEST/ \
  --sample-sheet /data/coredata/genomics/hiseq/150827_SN1046_0229_BC6WPEACXX/SampleSheet.csv \
  --use-bases-mask Y*,I8n*,Y* \
  --barcode-mismatches 2 -l DEBUG &

Sample sheet Format is below:

[Data],,,,,,,,
Lane,Sample_ID,Sample_Name,Sample_Plate,Sample_Well,Index_ID,Index,Sample_Project,Description
1,DPWGS-00088,DPWGS-00088,,,,NoIndex,DatePalm_-_Date_Fruits,
2,DPWGS-00089,DPWGS-00089,,,,NoIndex,DatePalm_-_Date_Fruits,
3,DPWGS-00072,DPWGS-00072,,,,NoIndex,DatePalm_-_Date_Fruits,
4,DPWGS-00078,DPWGS-00078,,,,NoIndex,DatePalm_-_Date_Fruits,
5,DPWGS-00247,DPWGS-00247,,,,AGTACAAG,DatePalm_-_Date_Fruits,
5,DPWGS-00267,DPWGS-00267,,,,GACTAGTA,DatePalm_-_Date_Fruits,
6,DPWGS-00263,DPWGS-00263,,,,AGTCACTA,DatePalm_-_Date_Fruits,
6,DPWGS-00268,DPWGS-00268,,,,CAATGGAA,DatePalm_-_Date_Fruits,
7,32,32,,,,GACTAGTA,DatePalm_-_Date_Fruits,
7,DPWGS-00079,DPWGS-00079,,,,CTGAGCCA,DatePalm_-_Date_Fruits,
8,31,31,,,,NoIndex,DatePalm_-_Date_Fruits,

Status Report are below

Flowcell Summary

Clusters (Raw)  Clusters(PF)   Yield (MBases)
2,243,033,353   1,710,124,620  342,025

Lane Summary

Lane   PF Clusters   % of the  % Perfect  % One mismatch  Yield (Mbases)   % PF      % >= Q30  Mean Quality
                     lane      barcode    barcode                          Clusters  bases     Score
1      0                                                  46,813           74.81     75.65     33.00
2      0                                                  40,294           64.81     72.67     32.12
3      0                                                  46,675           91.71     87.04     35.26
4      0                                                  13,543           26.86     53.72     27.30
5      244,751,056   100.00    97.46      1.88            48,950           84.95     79.86     33.46
6      237,309,355   100.00    72.61      25.73           47,462           91.76     87.45     35.40
7      243,656,285   100.00    92.56      6.93            48,731           90.96     86.39     35.19
8      0                                                  49,557           83.13     80.02     33.

[BioRyder]

hello Ido Tamir,

Thank you for your reply.Yes I have removed Noindex into empty while converting the data.Report file is based on that changes.

[BioRyder]

NoIndex changed to empty while running the script

[BioRyder]

Hi All,

We have identified the problem.The above mentioned problem is happened due to File format of linux Server. Bcl2fast Version is working properly in XFS file system. But If we are using gpfs file system in Linux server,bcl2fastq V2 is generating partial out put file, missing R1 or both R1 and R2. We have contacted illumina and informed the same. They are internally checking the issue of Bcl2fast with gpfs file system.

Answer 1

[dbrawand]

I have a similar problem when writing on a NFS share. Some of the fastq files get scrambled at the end or are incomplete (gzip CRC fails on decompression)

Setting the writing threads to 1 has solved this problem in my case (option -w 1).

Answer 2

[Ido Tamir]

a) [e-mail] is the best adress for this question

b) dont write noIndex, simply leave the index field empty

c) how can you have data for lane 4?

Answer 3

[BioRyder]

Hi All,

We have identified the problem.The above mentioned problem is happened due to File format of linux Server. Bcl2fast Version is working properly in XFS file system. But If we are using gpfs file system in Linux server,bcl2fastq V2 is generating partial out put file, missing R1 or both R1 and R2. We have contacted illumina and informed the same. They are internally checking the issue of Bcl2fast with gpfs file system.