# Error with bowtie2: Saw ASCII character 10 but expected 33-based Phred qual

**URL:** <https://forum.biobakery.org/t/error-with-bowtie2-saw-ascii-character-10-but-expected-33-based-phred-qual/6867>\
**Category:** KneadData\
**Created:** [March 29, 2024, 9:03am UTC](https://forum.biobakery.org/t/error-with-bowtie2-saw-ascii-character-10-but-expected-33-based-phred-qual/6867 "2024-03-29T09:03:36Z")\
**Posts on this page:** 1\
**Page:** 1

<div class="post-metadata">

**Author:** ![mojiefei](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/mojiefei/32/2802_2.png) [@mojiefei](https://forum.biobakery.org/u/mojiefei)\
**Post date:** [March 29, 2024, 9:03am UTC](https://forum.biobakery.org/t/error-with-bowtie2-saw-ascii-character-10-but-expected-33-based-phred-qual/6867/1 "2024-03-29T09:03:36Z")

</div>

_kneaddata v0.10.0_

**Issue 1:**  
I installed _kneaddata v0.10.0_ through conda. When I used one database as an index for _bowtie2_, everything ran fine except deleting the intermediate files containing the index name. Following are my scripts.

```auto
$ kneaddata -i /home/gfz3/mojf/rawdata/B11_1.fq.gz \
-i /home/gfz3/mojf/rawdata/B11_2.fq.gz \
-o /hdd/mojf/output/kneaddata \
-t 20 -p 20 \
-db /database/kneaddata_db/invertebrate_db/INVERTEBRATE \
--run-fastqc-start --run-fastqc-end \
--trimmomatic /home/gfz3/miniconda3/envs/kneaddata/bin/Trimmomatic-0.39 \
--trimmomatic-options "SLIDINGWINDOW:4:15 MINLEN:36" \
--remove-intermediate-output

```

**Issue 2:**  
When I used two databases as indexes, _bowtie2_ ran with an error. The error message I got is as follows.

```auto
Saw ASCII character 10 but expected 33-based Phred qual.
terminate called after throwing an instance of 'int' Aborted (core dumped)
(ERR): bowtie2-align exited with value 134

```

I just added the second _-db_ to the previous scripts and didn’t change anything else. Following are my scripts.

```auto
$ kneaddata -i /home/gfz3/mojf/rawdata/B11_1.fq.gz \
-i /home/gfz3/mojf/rawdata/B11_2.fq.gz \
-o /hdd/mojf/output/kneaddata \
-t 20 -p 20 \
-db /database/kneaddata_db/invertebrate_db/INVERTEBRATE \
-db /database/kneaddata_db/vertebrate_db/VERTEBRATE \
--run-fastqc-start --run-fastqc-end \
--trimmomatic /home/gfz3/miniconda3/envs/kneaddata/bin/Trimmomatic-0.39 \
--trimmomatic-options "SLIDINGWINDOW:4:15 MINLEN:36" \
--remove-intermediate-output

```

The first 8 lines of _B11\_1.fq.gz_ are as follows.

```auto
@LH00391:13:22F2NMLT3:5:1101:8055:1048 1:N:0:GAGCAAGATC+AAGTTACGGA
CNGACAACAACAGTGAGAAAAAAATAAAATAGCAGCCAAGAAGTACCGGAATACTACATCCATTGGTATCCAAGACGTCTGCGTGCAAGCACTTAGAAGAACGGGGGCAAAGTTTGAAAATATGGCCAAGTTCGATACCCTTGGGGAACT
+
I#IIIII9III-III-I--999999---------9---9-9---9--9999------9-9-9---9-9I9I99999-I-II99999-I9---I9999999--99--9-9-999-99999II9999--9-9-9I9999-99I--9--I9--
@LH00391:13:22F2NMLT3:5:1101:12326:1048 1:N:0:GAGCAAGATC+AAGTTGCGGA
TNAGAATCACGAGCAGCTAGCAAAACACGAAATAATAACAGCCGGAAATCCCAGGACCAAGGATGGACGCCAGCTGAGTCAAAGTCCTTCCATTCAACTACCCGCCGCCGGTCTGGTGACGCATAGCATCCAAAGTTACCAATATGAGCA
+
I#IIIIII9IIIIIIIIIIIIII-I-9--II-II99-9-9--9-9999-9999----99--99-I99-99-III---9--9I9999-9I----999---9-9----99-I-9--9-I-I-9-999----999-999999999-9-99999

```

The first 2 lines of _invertebrate.fasta_ used for indexing are as follows. The 6 files with the suffix _.bt2_ as the output of index were complete.

```auto
>NC_004354.4 Drosophila melanogaster chromosome X
GAATTCGTCAGAAATGAgctaaacaaatttaaatcattaaatgcGAGCGGCGAATCCGGAAACAGCAACTTCAAACCAGT

```

**I would like to know why the above two issues arise, especially the second one, and how to solve them. Looking forward to the solutions.**