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ValueError: Mixed RNA/DNA found. #61

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shaos1996 opened this issue Nov 14, 2024 · 15 comments
Open

ValueError: Mixed RNA/DNA found. #61

shaos1996 opened this issue Nov 14, 2024 · 15 comments

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@shaos1996
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I analyzed the intact seq extracted by LTR_retriever with the command 'TEsorter acau.intact.fa -db rexdb-plant -p 36 -rule 70-30-80'. After splitting the sequence into 72 pieces and translating it, an error was reported. Please tell me what is going on with this issue.

BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
BiopythonWarning,
2024-11-14 13:25:00,730 -INFO- translating ./tmp-b151e46e-a248-11ef-993b-ac1f6bc1b736/chunk.72.fasta in six frames
multiprocessing.pool.RemoteTraceback:
"""
Traceback (most recent call last):
File "/public1/user/shaoshao/software/miniconda3/lib/python3.7/multiprocessing/pool.py", line 121, in worker
result = (True, func(*args, **kwds))
File "/public1/user/shaoshao/software/miniconda3/lib/python3.7/multiprocessing/pool.py", line 44, in mapstar
return list(map(*args))
File "/public1/user/shaoshao/software/miniconda3/lib/python3.7/site-packages/TEsorter-1.4.7-py3.7.egg/TEsorter/app.py", line 1100, in _translate
return translate(inSeq, overwrite=overwrite)
File "/public1/user/shaoshao/software/miniconda3/lib/python3.7/site-packages/TEsorter-1.4.7-py3.7.egg/TEsorter/app.py", line 1080, in translate
six_frame_translate(inSeq, fp)
File "/public1/user/shaoshao/software/miniconda3/lib/python3.7/site-packages/TEsorter-1.4.7-py3.7.egg/TEsorter/modules/translate_seq.py", line 10, in six_frame_translate
for seq, suffix0 in zip([rc.seq, rc.seq.reverse_complement()], ['aa', 'rev_aa']):
File "/public1/user/shaoshao/software/python37/lib/python3.7/site-packages/Bio/Seq.py", line 1911, in reverse_complement
return self.complement()[::-1]
File "/public1/user/shaoshao/software/python37/lib/python3.7/site-packages/Bio/Seq.py", line 1847, in complement
raise ValueError("Mixed RNA/DNA found")
ValueError: Mixed RNA/DNA found
@zhangrengang
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@shaos1996 Please check: are both T and U in your fasta file?

@shaos1996
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shaos1996 commented Nov 14, 2024 •
edited
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No sequence containing U. and I tested it on 5 genomes and only one did not report an error.

@shaos1996
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This is the output of the bug, it seems that some standard output and sequences are connected together.

Debug - self._data content: b'GTCTATGTGTATGTGTTTGATTGTTTCTCTCTTGCTTAGGAACTAGGACATGTGCATGACGAAATGTTAAATGCTTAGGAACCTAAGAACATCTTTGCTATAAGGGGTTAACTCCTAAAGGAATTGCAACCAAGCATAACCTTTCTCTCACTCTCTTTCTCTCTTTTAAGGATACTTAAAGCACCCAAAAGAACTTAAAGTAATTAAGTATTTTCCGCTGCATGTTTATCATGAAAATACCTAGATGCATGACAre-organizeacau.fa.retriever.all.scnacau.fa.retriever.all.scntotal678715,0withoutchr,11402discarded,667313retained'/public1/user/shaoshao/software/miniconda3/lib/python3.7/site-packages/Bio/Seq.py:2750: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
BiopythonWarning,
/public1/user/shaoshao/software/miniconda3/lib/python3.7/site-packages/Bio/Seq.py:1639: BiopythonDeprecationWarning: seq.reverse_complement() will change in the near future to always return DNA nucleotides only. Please use

seq.reverse_complement_rna()

@zhangrengang
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@shaos1996 Can you send me your fasta file to debug?

@shaos1996
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I found the problem. It was that nohup was used in LTR_retriever.py get_full_seqs genome.fa > intact_ltr.fa. Remove nohup and it works fine.

@zhangrengang
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Yes, nohup rediects strandard error to strandard output.

@yaoxkkkkk
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yaoxkkkkk commented Nov 28, 2024 •
edited
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Hi I met the same error, here is the log:

/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py:2334: BiopythonWarning: Partial codon, len(sequence) not a multiple of three. Explicitly trim the sequence or add trailing N before translation. This may become an error in future.
  warnings.warn(
2024-11-28 23:52:31,498 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.21.fasta` in six frames
2024-11-28 23:52:34,762 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.22.fasta` in six frames
2024-11-28 23:52:34,942 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.23.fasta` in six frames
2024-11-28 23:52:35,079 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.24.fasta` in six frames
2024-11-28 23:52:35,109 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.25.fasta` in six frames
2024-11-28 23:52:35,110 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.26.fasta` in six frames
2024-11-28 23:52:35,161 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.27.fasta` in six frames
2024-11-28 23:52:35,168 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.28.fasta` in six frames
2024-11-28 23:52:35,169 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.29.fasta` in six frames
2024-11-28 23:52:35,215 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.30.fasta` in six frames
2024-11-28 23:52:35,244 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.31.fasta` in six frames
2024-11-28 23:52:35,248 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.32.fasta` in six frames
2024-11-28 23:52:35,260 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.33.fasta` in six frames
2024-11-28 23:52:35,265 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.34.fasta` in six frames
2024-11-28 23:52:35,272 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.35.fasta` in six frames
2024-11-28 23:52:35,292 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.36.fasta` in six frames
2024-11-28 23:52:35,294 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.37.fasta` in six frames
2024-11-28 23:52:35,296 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.38.fasta` in six frames
2024-11-28 23:52:35,324 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.39.fasta` in six frames
2024-11-28 23:52:35,332 -INFO- translating `./tmp-c38fe47e-ada0-11ef-9b74-7c8ae1d406bf/chunk.40.fasta` in six frames
multiprocessing.pool.RemoteTraceback: 
"""
Traceback (most recent call last):
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/multiprocessing/pool.py", line 125, in worker
    result = (True, func(*args, **kwds))
                    ^^^^^^^^^^^^^^^^^^^
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/multiprocessing/pool.py", line 48, in mapstar
    return list(map(*args))
           ^^^^^^^^^^^^^^^^
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/TEsorter/app.py", line 1100, in _translate
    return translate(inSeq, overwrite=overwrite)
           ^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/TEsorter/app.py", line 1080, in translate
    six_frame_translate(inSeq, fp)
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/TEsorter/modules/translate_seq.py", line 10, in six_frame_translate
    for seq, suffix0 in zip([rc.seq, rc.seq.reverse_complement()], ['aa', 'rev_aa']):
                                     ^^^^^^^^^^^^^^^^^^^^^^^^^^^
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py", line 825, in reverse_complement
    return self.complement()[::-1]
           ^^^^^^^^^^^^^^^^^
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/Bio/Seq.py", line 761, in complement
    raise ValueError("Mixed RNA/DNA found")
ValueError: Mixed RNA/DNA found
"""

The above exception was the direct cause of the following exception:

Traceback (most recent call last):
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/bin/TEsorter", line 10, in <module>
    sys.exit(main())
             ^^^^^^
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/TEsorter/app.py", line 1265, in main
    pipeline(Args())
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/TEsorter/app.py", line 257, in pipeline
    gff, geneSeq = LTRlibAnn(
                   ^^^^^^^^^^
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/TEsorter/app.py", line 1165, in LTRlibAnn
    chunk_files = hmmscan_pp(ltrlib, hmmdb=hmmdb, hmmout=domtbl, tmpdir=tmpdir, 
                  ^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/TEsorter/app.py", line 1118, in hmmscan_pp
    chunk_files = list(pool_func(_translate, iterable, processors=processors))
                  ^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/site-packages/TEsorter/modules/RunCmdsMP.py", line 347, in pool_func
    for returned in pool_map(func, iterable, **kargs):
                    ^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/multiprocessing/pool.py", line 367, in map
    return self._map_async(func, iterable, mapstar, chunksize).get()
           ^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^
  File "/dssg/home/acct-jiang.lu/jiang.lu-user1/mambaforge/envs/tesorter/lib/python3.11/multiprocessing/pool.py", line 774, in get
    raise self._value
ValueError: Mixed RNA/DNA found

I have checked the input TE lib file by seqkit, it returned:

�[33m[WARN]�[0m it's already DNA, no need to convert

I can't find nohup in related script, how can I resolve it?

@zhangrengang
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zhangrengang commented Nov 29, 2024 •
edited
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@yaoxkkkkk Please check the input TE lib file by cat your_TE_lib.fasta | grep -v ">" | grep -Po "[^ATCG]" -i.

@yaoxkkkkk
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yaoxkkkkk commented Nov 29, 2024 •
edited
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It returns lots of non-ATCG:

cat XZ.mod.fa.mod.EDTA.TElib.fa | grep -v ">" | grep -Po "[^ATCG]" -i
w
w
r
w
s
w
y
r
k
r
w
y
s
w
r
y
y
r
s
...

I will check my TE lib again, thank you for your reply!

@yaoxkkkkk
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I checked the file I merged, these non-ATCG characters are from athrep.updated.nonredun.fasta provided by EDTA, is there anything that I missed for using this file?

@yaoxkkkkk
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I realise they may IUPAC code, TEsorter can deal with that?

@yaoxkkkkk
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And there are lots of X exits in the file, does it matter?
image

@zhangrengang
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Are these from the raw genome sequences? Are there some U characters? ValueError: Mixed RNA/DNA found is recognized by biopython. biopython can deal with IUPAC code. Check by cat XZ.mod.fa.mod.EDTA.TElib.fa | grep -v ">" | grep -Po "[^ATCG]" -i | sort |uniq -c

@yaoxkkkkk
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Yes it has, but seqkit fail to recognize them :(

cat XZ.mod.fa.mod.EDTA.TElib.mod.fa | grep -v ">" | grep -Po "[^ATCG]" -i | sort |uniq -c
    735 B
   1341 D
    111 E
     44 F
   1378 H
    103 I
  55290 K
     63 L
  65863 M
 168927 N
     81 O
     46 P
  22936 R
  36771 S
     39 U
    786 V
  75432 W
  36236 X
  23220 Y

@zhangrengang
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Some characters (e.g. E, F) are not DNA codes, and need to be checked, deleted or replaced.

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@zhangrengang @yaoxkkkkk @shaos1996