The analysis indicators for Smart-seq2 seem abnormal.

[Barry Huotari]

STAR --runThreadN 16 \
     --genomeDir sheep/geneome/STAR/ \
     --runDirPerm All_RWX \
     --readFilesCommand zcat \
         --outSAMtype None \
         --soloType SmartSeq \
         --readFilesManifest s1.mapfile \
     --soloUMIdedup Exact --soloStrand Unstranded \
         --limitOutSJcollapsed 10000000 --soloCellFilter None \
     --soloFeatures Gene GeneFull --soloOutFileNames output/ features.tsv barcodes.tsv matrix.mtx --outReadsUnmapped Fastx
     

cat Barcodes.stats
noNoAdapter 0
noNoUMI 0
noNoCB 0
noNinCB 0
noNinUMI 0
noUMIhomopolymer 0
noNoWLmatch 0
noTooManyMM 0
noTooManyWLmatches 0
yesWLmatchExact 20223089
yesOneWLmatchWithMM 0
yesMultWLmatchWithMM 0
GeneFull
cat Summary.csv
Number of Reads,20223089
Reads With Valid Barcodes,1
Sequencing Saturation,0.184216
Q30 Bases in RNA read,0.905109
Reads Mapped to Genome: Unique+Multiple,0.00719257
Reads Mapped to Genome: Unique,0.000716359
Reads Mapped to GeneFull: Unique+Multiple GeneFull,0.00217988
Reads Mapped to GeneFull: Unique GeneFull,0.00217988
cat Features.stats
noUnmapped 20077633
noNoFeature 82069
MultiFeature 19303
subMultiFeatureMultiGenomic 18407
noTooManyWLmatches 0
noMMtoWLwithoutExact 0
yesWLmatch 44084
yessubWLmatchExact 44084
yessubWLmatch_UniqueFeature 44084
yesCellBarcodes 1
yesUMIs 35963

Gene
cat Summary.csv
Number of Reads,20223089
Reads With Valid Barcodes,1
Sequencing Saturation,0.2
Q30 Bases in RNA read,0.905109
Reads Mapped to Genome: Unique+Multiple,0.00719257
Reads Mapped to Genome: Unique,0.000716359
Reads Mapped to Gene: Unique+Multiple Gene,2.47242e-07
Reads Mapped to Gene: Unique Gene,2.47242e-07
cat Features.stats
noUnmapped 20077633
noNoFeature 145451
MultiFeature 0
subMultiFeatureMultiGenomic 0
noTooManyWLmatches 0
noMMtoWLwithoutExact 0
yesWLmatch 5
yessubWLmatchExact 5
yessubWLmatch_UniqueFeature 5
yesCellBarcodes 1
yesUMIs 4

I am comparing sheep genomes, and I feel that the Reads Mapped to Genome: Unique is a bit low. Is this normal?

[Alexander Dobin]

Something went very wrong in this run, I would recommend mapping a few reads from one file without single cell options.