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Report

The second file is prefix.lima.report, a tab-separated file about each ZMW, unfiltered. This report contains any information necessary to investigate the demultiplexing process and the underlying data. A single row contains all reads of a single ZMW. For --per-read, each row contains one subread and ZMWs might span multiple rows.

Lima reports different information for four basic entities First, FirstAlt, Combined, SecondCandidate, SecondAlt.

  • First is the barcode with the highest mean score.
  • FirstAlt is the barcode with the next highest mean score to First.
  • SecondCandidate is the barcode that in combination with First has the highest mean score.
  • SecondAlt is the barcode with the next highest mean score to SecondCandidate.
  • Combined is the combination of First and SecondCandidate, if the call has been deemed to be different, otherwise this entity is a copy of First.

Information about each entity:

  • Idx: Barcode index, 0-based. IdxCombined equals IdxSecondCandidate if different or IdxFirst if same.
  • Idx*Named: Barcode name.
  • Score: Mean score, omitting missing barcode regions.
  • Scores: List of scores for each barcode region.
  • Clips: List of clipping positions, leading and trailing alternating.

Additional columns:

  • ZMW: Movie name and the ZMW hole number.
  • ScoreCombinedFirst: The mean barcode score for the first barcode of Combined.
  • ScoreCombinedSecond: The mean barcode score for the second barcode of Combined.
  • ReadLengths: List of read lengths.
  • ScoreLead: The mean absolute element-wise score difference of the best to alternative pair.
  • SignalIncrease: Difference of ScoreFirst to a combination of ScoreFirst and ScoreSecondCandidate.
  • NumPasses: Number of full passes.
  • NumAdapters: Number of adapters.
  • IdxsCombined: The sequence of barcode indices IdxFirst and IdxCombined along the ZMW.
  • PassedFilters: 1, if reads of this ZMW passed user-provided filters and records were written to BAM, otherwise 0.
  • IdxLowest & IdxHighest: The sorted IdxFirst and IdxCombined. Sorting deactivated with --keep-idx-order.
  • IdxLowestNamed & IdxHighestNamed: The sorted IdxFirstNamed and IdxCombinedNamed. Sorting deactivated with -keep-idx-order.

Side note:

  • ScoreCombined is also referred to as Mean Score, Barcode Score, or Barcode Quality, depending who you are talking to.

Missing data: An individual score with -1 indicates that a leading or trailing adapter is missing. This is irrelevant for CCS reads, as they are full length and flanked by adapters. If there is no barcode region to call, fields are filled with a 0, -1, or - placeholders; please see example.

Example data:

$ head prefix.lima.report | column -t
ZMW                           IdxFirst  IdxCombined  IdxSecondCandidate  IdxFirstAlt  ScoreFirst  ScoreCombined  ScoreSecondCandidate  ScoreFirstAlt  ScoresFirst                      ScoresCombined                   ScoresSecondCandidate               ScoresFirstAlt                      ReadLengths                   IdxsCombined                                   ClipsFirst                                  ClipsCombined                               ClipsSecondCandidate                      ClipsFirstAlt                             ScoreLead  SignalIncrease  NumPasses  NumAdapters  PassedFilters  IdxFirstNamed  IdxCombinedNamed  IdxSecondCandidateNamed  IdxFirstAltNamed  IdxLowest  IdxHighest  IdxLowestNamed  IdxHighestNamed
m54007_170603_012109/4522478  366       366          136                 136          24          24             23                    23             -1,37,37,25,14,12,20,-1          -1,37,37,25,14,12,20,-1          -1,21,25,25,31,9,31,-1              -1,21,25,25,31,9,31,-1              1582,2392,2656,2243           -1,366,366,366,366,366,366,-1                  0,1561,19,2380,12,2654,18,2243              0,1561,19,2380,12,2654,18,2243              0,1574,14,2372,19,2651,13,2243            0,1574,14,2372,19,2651,13,2243            9          4               2          3            0              bc1367         bc1367            bc1137                   bc1137            366        366         bc1367          bc1367
m54007_170603_012109/4588251  265       265          1                   14           67          67             17                    56             -1,65,70,-1                      -1,65,70,-1                      -1,15,20,-1                         -1,54,59,-1                         1471,1698                     -1,265,265,-1                                  0,1456,17,1698                              0,1456,17,1698                              0,1453,16,1698                            0,1453,20,1698                            11         0               0          1            1              bc1266         bc1266            bc1002                   bc1015            265        265         bc1266          bc1266
m54007_170603_012109/4653426  213       213          325                 278          18          18             17                    18             -1,32,12,6,6,32,21,6,6,12,48,-1  -1,32,12,6,6,32,21,6,6,12,48,-1  -1,20,20,14,25,14,12,6,14,31,20,-1  -1,25,25,12,14,18,15,9,12,25,25,-1  715,2326,2267,2315,2332,1587  -1,213,213,213,213,213,213,213,213,213,213,-1  0,697,2,2325,1,2250,16,2314,1,2330,19,1587  0,697,2,2325,1,2250,16,2314,1,2330,19,1587  0,709,6,2321,4,2262,2,2314,5,2327,6,1587  0,711,4,2324,5,2264,6,2310,2,2328,4,1587  9          6               4          5            0              bc1214         bc1214            bc1326                   bc1279            213        213         bc1214          bc1214
m54007_170603_012109/4719449  -1        -1           -1                  -1           -1          -1             -1                    -1             -1                               -1                               -1                                  -1                                  292                           -                                              0                                           0                                           0                                         0                                         -1         -1              0          0            0              -1             -1                -1                       -1                -1         -1          -1              -1

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