RLMIDs vs GSMIDs

[Anthony.287]

Hello all!
I'm finally starting to get the hang of sfffile, and am finding some things that I can't explain. For instance, one particular region (a pool of 4 RLMID tagged rapid libraries) on the most recent PTP had 43,949 reads. 43,792 of those got sorted in sff's based on the RLMIDs. Out of curiosity, I sorted using the GSMIDs, and ~15,000 were sorted then. What could be happening?

[[email protected] ~]$ sfffile -s GSMIDs /data/R_2012_11_29_16_12_43_FLX05100626_Administrator_Shotgun/sff/HYPJBHH12_Shotgun.sff
Reading the input SFF file(s)...
Generating the split SFF file(s)...
MID1: 3 reads written into the SFF file.
MID2: 0 reads found.
MID3: 13 reads written into the SFF file.
MID4: 10898 reads written into the SFF file.
MID5: 4660 reads written into the SFF file.
MID6: 0 reads found.
MID7: 0 reads found.
MID8: 0 reads found.
MID9: 1 reads written into the SFF file.
MID10: 0 reads found.
MID11: 0 reads found.
MID12: 0 reads found.
MID13: 0 reads found.
MID14: 0 reads found.
[[email protected] ~]$ sfffile -s RLMIDs /data/R_2012_11_29_16_12_43_FLX05100626_Administrator_Shotgun/sff/HYPJBHH12_Shotgun.sff
Reading the input SFF file(s)...
Generating the split SFF file(s)...
RL1: 24344 reads written into the SFF file.
RL2: 11022 reads written into the SFF file.
RL3: 3692 reads written into the SFF file.
RL4: 4734 reads written into the SFF file.
RL5: 0 reads found.
RL6: 1 reads written into the SFF file.
RL7: 0 reads found.
RL8: 0 reads found.
RL9: 0 reads found.
RL10: 0 reads found.
RL11: 0 reads found.
RL12: 0 reads found.


Can the RLMIDs be misinterpreted as GSMIDs, and vice-versa? Any ideas?

Thank you!

[flxlex]

Yes, presumably because by default, up to two mismatches are allowed between the MID and the read. If you try allow only for one error, this should not happen. Either copy the Installation_path/454/config/MIDConfig.parse file to a new location and edit it:

mid = "MID4", "AGCACTGTAG", 2;

becomes

mid = "MID4", "AGCACTGTAG", 0;

Then run

sfffile -s GSMIDs -mcf modified_MIDConfig.parse /data/R_2012_11_29_16_12_43_FLX05100626_Administrator_Shotgun/sff/HYPJBHH12_Shotgun.sff

Or to indicate specific MIDs and allow no errors (adding '/0') run

sfffile -s GSMIDs:mid4/0,mid5/[email protected]/data/R_2012_11_29_16_12_43_FLX05100626_Administrator_Shotgun/sff/HYPJBHH12_Shotgun.sff
sfffile -s RLMIDs:mid1/0,mid2/0,mid3/0,mid4/[email protected]/data/R_2012_11_29_16_12_43_FLX05100626_Administrator_Shotgun/sff/HYPJBHH12_Shotgun.sff