<p>As @terdon mentions, it's certainly possible to have an <code>A</code> aligned on both strands. If a read has a mismatch at the location of interest and that mismatch is the complement of the reference base, then you are likely to see <code>A</code> in both strands. However, it is very unlikely you will see the same base <em>at high coverage</em> in both strands. For example, if your total coverage is 55, and you see 50 <code>A</code>s on the + strand and 50 <code>A</code>s on the - strand, it's likely that the pileup object is reporting the reverse complement sequence of the reads that align to the - strand for your convenience.</p>

CC BY-SA 4.0
2019-03-26T18:04:26.853Z
2019-03-26T18:04:26.853Z
Daniel Standage
<p>Is it possible that on the same <a href="https://pysam.readthedocs.io/en/latest/api.html?highlight=pileupcolumn#pysam.PileupColumn" rel="nofollow noreferrer">pileupcolumn</a> object (i.e a specific aligned base) I find the same base but aligned to different strands? For example, can I find a coverage of 50 'A's that align to strand '+', and on the same time 50 'A's that align to strand '-' within the same index? I expect that if 'A' is aligned to strand '+', I will find 'T' aligned to strand '-'.</p>

CC BY-SA 4.0
2019-03-26T14:55:16.873Z
Please [edit] your question and clarify what you are doing. What code are you running? On what input? Anything is possible, including mismatched bases either in the real data or because of a sequencing error, but we can't know if you don't give us more details.
terdon
<python><pysam>
Aligned base strand in pysam `pileupcolumn`
# Aligned base strand in pysam `pileupcolumn`

(Asked by: username_0 on 2019-03-26)

Is it possible that on the same [pileupcolumn](https://pysam.readthedocs.io/en/latest/api.html?highlight=pileupcolumn#pysam.PileupColumn) object (i.e a specific aligned base) I find the same base but aligned to different strands? For example, can I find a coverage of 50 'A's that align to strand '+', and on the same time 50 'A's that align to strand '-' within the same index? I expect that if 'A' is aligned to strand '+', I will find 'T' aligned to strand '-'.

**Tags:** python, pysam
> **Comments:**
> **username_1**: Please [edit] your question and clarify what you are doing. What code are you running? On what input? Anything is possible, including mismatched bases either in the real data or because of a sequencing error, but we can't know if you don't give us more details.  

---

## 1 answers

### Answer by username_2 on 2019-03-26. Score: 1

As @terdon mentions, it's certainly possible to have an `A` aligned on both strands. If a read has a mismatch at the location of interest and that mismatch is the complement of the reference base, then you are likely to see `A` in both strands. However, it is very unlikely you will see the same base *at high coverage* in both strands. For example, if your total coverage is 55, and you see 50 `A`s on the + strand and 50 `A`s on the - strand, it's likely that the pileup object is reporting the reverse complement sequence of the reads that align to the - strand for your convenience.


---

