some questions on cuffmerge and samtools flagstat output

i19870503

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some questions on cuffmerge and samtools flagstat output

09-11-2014, 06:41 PM

1.RNAseq is solid data mapped with bioscope, and I add XA tag manually to run cufflinks.
when I use cuffmerge , it shows:Warning: couldn't find fasta record for 'chr1'! and so on.
but I got merge.gtf, is there any different with normal output.

2.I check the bam file with samtools flagstat, but I think there is something wrong
54196276 + 0 in total (QC-passed reads + QC-failed reads)
0 + 0 duplicates
54196276 + 0 mapped (100.00%:-nan%)
0 + 0 paired in sequencing
0 + 0 read1
0 + 0 read2
0 + 0 properly paired (-nan%:-nan%)
0 + 0 with itself and mate mapped
0 + 0 singletons (-nan%:-nan%)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)

and also the samtools view output is here, did that mapped and sort correctly?
@HD VN:1.0 SO:coordinate
@SQ SN:chr1 LN:295529705 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:2cf1f3b70031dc473022a5a5893e8edb
@SQ SN:chr2 LN:140133492 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:b709bf0683d1abdca4f6236a45524577
@SQ SN:chr3 LN:123599780 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:2e896d4cc99e908405eb690a71467e37
@SQ SN:chr4 LN:136254946 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:32ceede4917f4ea64a7ab93040211d21
@SQ SN:chr5 LN:100516970 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:50bd953e6de61fb729390a122f830653
@SQ SN:chr6 LN:123305171 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:2ed43939ee3490a30cbe40e96cf33289
@SQ SN:chr7 LN:136409062 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:57ed74ddd703b537d8fece561887bcf4
@SQ SN:chr8 LN:119985671 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:deadaedc161e58bf8c8d6b3ea1c25eba
@SQ SN:chr9 LN:132468591 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:89eae5d26c774408170957bb006efbbd
@SQ SN:chr10 LN:66736929 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:171e7a3f72ba195f457720406be3604b
@SQ SN:chr11 LN:79814395 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:feeab3090c18a3cab9547f99d35052a8
@SQ SN:chr12 LN:57431344 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:f3309f09ea225f9a191b5d0d9eee961d
@SQ SN:chr13 LN:145235301 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:3a4b8ac5120541c225eb870fa4a2b501
@SQ SN:chr14 LN:148510138 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:c607e774abadba7ed0b0185f18e3f9d5
@SQ SN:chr15 LN:134541103 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:08f26a29f88b39bff614cd6fbfa7d2ef
@SQ SN:chr16 LN:77435658 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:0f5873a922aa3327b4523def56cad5e6
@SQ SN:chr17 LN:64395339 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:ac533e777ca34fe996fdf58598266b98
@SQ SN:chr18 LN:54309914 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:1cc33aec4862e0d56e691e5dc5960501
@SQ SN:chrX LN:125871292 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:d09f454e7e0e413bdd5e65af386c581d
@SQ SN:chrM LN:16770 UR:file:/mnt/SolidPool/reference/genomes/susScr2/swine_susScr2_validated.fa M5:c32d2133c7327cc9e47776c5eed17977
@RG ID:20121105185046758 LB:50F SM:SL002_251012_FC1_PG2_F3
@PG ID:bioscope-genome-mapping VN:bioscope-1.2 CL:mapping.qual.pvalue=1; mapping.mismatch.penalty=-2.0; mapping.run.classic=false; mapping.qual.anchor.length=25; matching.max.hits=10; mapping.qual.filter.cutoff=0; mapping.classic.anchor.length=50; mapping.valid.adjacent=0; mapping.classic.mismatch=0; mapping.scheme.unmapped=25.2.0\:20; matching.use.iub.reference=false; mapping.qual.error.rate=0.2; mapping.qual.mismatch.level=3; mapping.qual.bvalue=1.0; read.length=50; mapping.scheme.repetitive=
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