Mixing
Total reads aligning to each reference within a bam file
Clash Royale CLAN TAG#URR8PPP
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I have two PCR amplicons that have been multiplexed and sequenced using the nanopore minion.
I have aligned the fastq reads using minimap2 with a reference file containing both amplicon sequences and generated a bam file that I have viewed using IGV.
I am looking for a way to generate some simple summary statistics.
In particular, is there a way to extract the total number of fastq reads aligning to each amplicon reference from the bam file?
alignment bam nanopore minion
edited 4 hours ago
Llopis
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asked 4 hours ago
CM3
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up vote
3
down vote
favorite
I have two PCR amplicons that have been multiplexed and sequenced using the nanopore minion.
I have aligned the fastq reads using minimap2 with a reference file containing both amplicon sequences and generated a bam file that I have viewed using IGV.
I am looking for a way to generate some simple summary statistics.
In particular, is there a way to extract the total number of fastq reads aligning to each amplicon reference from the bam file?
alignment bam nanopore minion
edited 4 hours ago
Llopis
2,5471628
asked 4 hours ago
CM3
161
New contributor
CM3 is a new contributor to this site. Take care in asking for clarification, commenting, and answering.
Check out our Code of Conduct.
add a comment |Â
up vote
3
down vote
favorite
up vote
3
down vote
favorite
I have two PCR amplicons that have been multiplexed and sequenced using the nanopore minion.
I have aligned the fastq reads using minimap2 with a reference file containing both amplicon sequences and generated a bam file that I have viewed using IGV.
I am looking for a way to generate some simple summary statistics.
In particular, is there a way to extract the total number of fastq reads aligning to each amplicon reference from the bam file?
alignment bam nanopore minion
edited 4 hours ago
Llopis
2,5471628
asked 4 hours ago
CM3
161
New contributor
CM3 is a new contributor to this site. Take care in asking for clarification, commenting, and answering.
Check out our Code of Conduct.
I have two PCR amplicons that have been multiplexed and sequenced using the nanopore minion.
I have aligned the fastq reads using minimap2 with a reference file containing both amplicon sequences and generated a bam file that I have viewed using IGV.
I am looking for a way to generate some simple summary statistics.
In particular, is there a way to extract the total number of fastq reads aligning to each amplicon reference from the bam file?
alignment bam nanopore minion
alignment bam nanopore minion
edited 4 hours ago
Llopis
2,5471628
asked 4 hours ago
CM3
161
New contributor
CM3 is a new contributor to this site. Take care in asking for clarification, commenting, and answering.
Check out our Code of Conduct.
edited 4 hours ago
Llopis
2,5471628
asked 4 hours ago
CM3
161
New contributor
CM3 is a new contributor to this site. Take care in asking for clarification, commenting, and answering.
Check out our Code of Conduct.
edited 4 hours ago
Llopis
2,5471628
edited 4 hours ago
Llopis
2,5471628
edited 4 hours ago
Llopis
2,5471628
2,5471628
asked 4 hours ago
CM3
161
New contributor
CM3 is a new contributor to this site. Take care in asking for clarification, commenting, and answering.
Check out our Code of Conduct.
asked 4 hours ago
CM3
161
asked 4 hours ago
CM3
161
161
New contributor
CM3 is a new contributor to this site. Take care in asking for clarification, commenting, and answering.
Check out our Code of Conduct.
New contributor
CM3 is a new contributor to this site. Take care in asking for clarification, commenting, and answering.
Check out our Code of Conduct.
CM3 is a new contributor to this site. Take care in asking for clarification, commenting, and answering.
Check out our Code of Conduct.
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3 Answers
3
active
oldest
votes
up vote
1
down vote
This can be done using
samtools flagstat your_bam_file
answered 4 hours ago
Ammar Sabir Cheema
513213
I think that flagstat counts the number of alignments in the bam file.
– Peter Menzel
3 hours ago
add a comment |Â
up vote
1
down vote
This one-liner below will work better for long reads than samtools flagstat in that it only counts the primary alignment for each read and samtools flagstat doesn't seem to calculate some stats for long reads. I have never seen samtools flagstat output stats on a per-reference basis, but am curious if so!
This answer filters out secondary and supplementary alignments for your reads (-F 2304) that have some alignment to both amplicon reference and just keep the best one. This might give a more accurate idea of how many reads of each amplicon are in the library.
samtools view -F 2304 myfile.bam | awk -F $'t' 'a[$1, $3]++ ENDfor (i in a) split (i, sep, SUBSEP); print sep[1], sep[2], a[i]' | uniq | awk 'print($2)' | uniq -c | sort -k1 -nr
adapted from this.
answered 4 hours ago
conchoecia
96019
add a comment |Â
up vote
1
down vote
The quick way to get the number of alignments on each reference is
samtools idxstats my_bam.bam
Number of reads on each reference is column 3. Although, as has been pointed out, this will give you the total number of alignments per reference, not the total number of reads (each read might give rise to more than one alignment). That said I do tend to us this as generally I'm after a rough approximation, rather than an accurate number.
In theory, only one alignment for each read should be marked as primary, so the following should give you what you need quickly and at low memory usage:
samtools view -bF 2304 my_bam.bam > primary_only.bam
samtools index primary_only.bam
samtools idxstats primary_only.bam
answered 1 hour ago
Ian Sudbery
1,656214
add a comment |Â
3 Answers
3
active
oldest
votes
3 Answers
3
active
oldest
votes
active
oldest
votes
active
oldest
votes
up vote
1
down vote
This can be done using
samtools flagstat your_bam_file
answered 4 hours ago
Ammar Sabir Cheema
513213
I think that flagstat counts the number of alignments in the bam file.
– Peter Menzel
3 hours ago
add a comment |Â
up vote
1
down vote
This can be done using
samtools flagstat your_bam_file
answered 4 hours ago
Ammar Sabir Cheema
513213
I think that flagstat counts the number of alignments in the bam file.
– Peter Menzel
3 hours ago
add a comment |Â
up vote
1
down vote
up vote
1
down vote
This can be done using
samtools flagstat your_bam_file
answered 4 hours ago
Ammar Sabir Cheema
513213
This can be done using
samtools flagstat your_bam_file
answered 4 hours ago
Ammar Sabir Cheema
513213
answered 4 hours ago
Ammar Sabir Cheema
513213
answered 4 hours ago
Ammar Sabir Cheema
513213
answered 4 hours ago
Ammar Sabir Cheema
513213
513213
I think that flagstat counts the number of alignments in the bam file.
– Peter Menzel
3 hours ago
add a comment |Â
I think that flagstat counts the number of alignments in the bam file.
– Peter Menzel
3 hours ago
I think that flagstat counts the number of alignments in the bam file.
– Peter Menzel
3 hours ago
I think that flagstat counts the number of alignments in the bam file.
– Peter Menzel
3 hours ago
add a comment |Â
up vote
1
down vote
This one-liner below will work better for long reads than samtools flagstat in that it only counts the primary alignment for each read and samtools flagstat doesn't seem to calculate some stats for long reads. I have never seen samtools flagstat output stats on a per-reference basis, but am curious if so!
This answer filters out secondary and supplementary alignments for your reads (-F 2304) that have some alignment to both amplicon reference and just keep the best one. This might give a more accurate idea of how many reads of each amplicon are in the library.
samtools view -F 2304 myfile.bam | awk -F $'t' 'a[$1, $3]++ ENDfor (i in a) split (i, sep, SUBSEP); print sep[1], sep[2], a[i]' | uniq | awk 'print($2)' | uniq -c | sort -k1 -nr
adapted from this.
answered 4 hours ago
conchoecia
96019
add a comment |Â
up vote
1
down vote
This one-liner below will work better for long reads than samtools flagstat in that it only counts the primary alignment for each read and samtools flagstat doesn't seem to calculate some stats for long reads. I have never seen samtools flagstat output stats on a per-reference basis, but am curious if so!
This answer filters out secondary and supplementary alignments for your reads (-F 2304) that have some alignment to both amplicon reference and just keep the best one. This might give a more accurate idea of how many reads of each amplicon are in the library.
samtools view -F 2304 myfile.bam | awk -F $'t' 'a[$1, $3]++ ENDfor (i in a) split (i, sep, SUBSEP); print sep[1], sep[2], a[i]' | uniq | awk 'print($2)' | uniq -c | sort -k1 -nr
adapted from this.
answered 4 hours ago
conchoecia
96019
add a comment |Â
up vote
1
down vote
up vote
1
down vote
This one-liner below will work better for long reads than samtools flagstat in that it only counts the primary alignment for each read and samtools flagstat doesn't seem to calculate some stats for long reads. I have never seen samtools flagstat output stats on a per-reference basis, but am curious if so!
This answer filters out secondary and supplementary alignments for your reads (-F 2304) that have some alignment to both amplicon reference and just keep the best one. This might give a more accurate idea of how many reads of each amplicon are in the library.
samtools view -F 2304 myfile.bam | awk -F $'t' 'a[$1, $3]++ ENDfor (i in a) split (i, sep, SUBSEP); print sep[1], sep[2], a[i]' | uniq | awk 'print($2)' | uniq -c | sort -k1 -nr
adapted from this.
answered 4 hours ago
conchoecia
96019
This one-liner below will work better for long reads than samtools flagstat in that it only counts the primary alignment for each read and samtools flagstat doesn't seem to calculate some stats for long reads. I have never seen samtools flagstat output stats on a per-reference basis, but am curious if so!
This answer filters out secondary and supplementary alignments for your reads (-F 2304) that have some alignment to both amplicon reference and just keep the best one. This might give a more accurate idea of how many reads of each amplicon are in the library.
samtools view -F 2304 myfile.bam | awk -F $'t' 'a[$1, $3]++ ENDfor (i in a) split (i, sep, SUBSEP); print sep[1], sep[2], a[i]' | uniq | awk 'print($2)' | uniq -c | sort -k1 -nr
adapted from this.
answered 4 hours ago
conchoecia
96019
answered 4 hours ago
conchoecia
96019
answered 4 hours ago
conchoecia
96019
answered 4 hours ago
conchoecia
96019
96019
add a comment |Â
add a comment |Â
up vote
1
down vote
The quick way to get the number of alignments on each reference is
samtools idxstats my_bam.bam
Number of reads on each reference is column 3. Although, as has been pointed out, this will give you the total number of alignments per reference, not the total number of reads (each read might give rise to more than one alignment). That said I do tend to us this as generally I'm after a rough approximation, rather than an accurate number.
In theory, only one alignment for each read should be marked as primary, so the following should give you what you need quickly and at low memory usage:
samtools view -bF 2304 my_bam.bam > primary_only.bam
samtools index primary_only.bam
samtools idxstats primary_only.bam
answered 1 hour ago
Ian Sudbery
1,656214
add a comment |Â
up vote
1
down vote
The quick way to get the number of alignments on each reference is
samtools idxstats my_bam.bam
Number of reads on each reference is column 3. Although, as has been pointed out, this will give you the total number of alignments per reference, not the total number of reads (each read might give rise to more than one alignment). That said I do tend to us this as generally I'm after a rough approximation, rather than an accurate number.
In theory, only one alignment for each read should be marked as primary, so the following should give you what you need quickly and at low memory usage:
samtools view -bF 2304 my_bam.bam > primary_only.bam
samtools index primary_only.bam
samtools idxstats primary_only.bam
answered 1 hour ago
Ian Sudbery
1,656214
add a comment |Â
up vote
1
down vote
up vote
1
down vote
The quick way to get the number of alignments on each reference is
samtools idxstats my_bam.bam
Number of reads on each reference is column 3. Although, as has been pointed out, this will give you the total number of alignments per reference, not the total number of reads (each read might give rise to more than one alignment). That said I do tend to us this as generally I'm after a rough approximation, rather than an accurate number.
In theory, only one alignment for each read should be marked as primary, so the following should give you what you need quickly and at low memory usage:
samtools view -bF 2304 my_bam.bam > primary_only.bam
samtools index primary_only.bam
samtools idxstats primary_only.bam
answered 1 hour ago
Ian Sudbery
1,656214
The quick way to get the number of alignments on each reference is
samtools idxstats my_bam.bam
Number of reads on each reference is column 3. Although, as has been pointed out, this will give you the total number of alignments per reference, not the total number of reads (each read might give rise to more than one alignment). That said I do tend to us this as generally I'm after a rough approximation, rather than an accurate number.
In theory, only one alignment for each read should be marked as primary, so the following should give you what you need quickly and at low memory usage:
samtools view -bF 2304 my_bam.bam > primary_only.bam
samtools index primary_only.bam
samtools idxstats primary_only.bam
answered 1 hour ago
Ian Sudbery
1,656214
answered 1 hour ago
Ian Sudbery
1,656214
answered 1 hour ago
Ian Sudbery
1,656214
answered 1 hour ago
Ian Sudbery
1,656214
1,656214
add a comment |Â
add a comment |Â
CM3 is a new contributor. Be nice, and check out our Code of Conduct.
CM3 is a new contributor. Be nice, and check out our Code of Conduct.
CM3 is a new contributor. Be nice, and check out our Code of Conduct.
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