What sequences should I use for ATAC-Seq adapter trimming using trimmomatic 0.36?

Question

[Gary]

Hi,

I want to use Trimmomatic 0.36 to trim ATAC-Seq adapter sequences. However, I am not sure which sequences I should use to trim our fastq files. Should I use the NexteraPE-PE.fa file provided by Trimmomatic 0.36 based on two Biostars threads below?

Biostars thread I: The only odd thing I see is that you trimmed for the TruSeq instead of the Nextera adapter.

ATAC-seq fragment length distribution

Biostars thread II: I chose Trimmomatic in the galaxy of my university (Illuminaclip Nextera pair end adapter) to trim my fastq file

How to treat the unpaired data after trimmomatic

By the way, I have tried (1) NexteraPE-PE.fa provided by Trimmomatic (the detail below), and (2) ATACseq.fa, including Ad1_noMX & Ad2.5_GGACTCCT primers we used (the detail below). Below are their commands and results. Could you help me? May you share the correct sequence file with me for ATAC-Seq adapter trimming using Trimmomatic? Thank you very much.

Trimmomatic commands & results I. using NexteraPE-PE.fa provided by Trimmomatic

gary > java -jar /Users/gary/Applications/Trimmomatic-0.36/trimmomatic-0.36.jar PE -threads 6 -trimlog ATACseqNexteraPE.txt -phred33 bulbul_R1.fastq.gz bulbul_R2.fastq.gz bulbul_R1_paired.fastq.gz bulbul_R1_unpaired.fastq.gz bulbul_R2_paired.fastq.gz bulbul_R2_unpaired.fastq.gz ILLUMINACLIP:/Users/gary/Applications/Trimmomatic-0.36/adapters/NexteraPE-PE.fa:2:30:10:8:true
TrimmomaticPE: Started with arguments:
 -threads 6 -trimlog ATACseqNexteraPE.txt -phred33 bulbul_R1.fastq.gz bulbul_R2.fastq.gz bulbul_R1_paired.fastq.gz bulbul_R1_unpaired.fastq.gz bulbul_R2_paired.fastq.gz bulbul_R2_unpaired.fastq.gz ILLUMINACLIP:/Users/gary/Applications/Trimmomatic-0.36/adapters/NexteraPE-PE.fa:2:30:10:8:true
Using PrefixPair: 'AGATGTGTATAAGAGACAG' and 'AGATGTGTATAAGAGACAG'
Using Long Clipping Sequence: 'GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAG'
Using Long Clipping Sequence: 'TCGTCGGCAGCGTCAGATGTGTATAAGAGACAG'
Using Long Clipping Sequence: 'CTGTCTCTTATACACATCTCCGAGCCCACGAGAC'
Using Long Clipping Sequence: 'CTGTCTCTTATACACATCTGACGCTGCCGACGA'
ILLUMINACLIP: Using 1 prefix pairs, 4 forward/reverse sequences, 0 forward only sequences, 0 reverse only sequences
Input Read Pairs: 26847985 Both Surviving: 26759022 (99.67%) Forward Only Surviving: 22244 (0.08%) Reverse Only Surviving: 64789 (0.24%) Dropped: 1930 (0.01%)
TrimmomaticPE: Completed successfully

II. Using ATACseq.fa, including Ad1_noMX & Ad2.5_GGACTCCT primers

gary > java -jar /Users/gary/Applications/Trimmomatic-0.36/trimmomatic-0.36.jar PE -threads 6 -trimlog ATACseq.txt -phred33 bulbul_R1.fastq.gz bulbul_R2.fastq.gz bulbul_R1_paired.fastq.gz bulbul_R1_unpaired.fastq.gz bulbul_R2_paired.fastq.gz bulbul_R2_unpaired.fastq.gz ILLUMINACLIP:/Users/gary/Applications/Trimmomatic-0.36/adapters/ATACseq.fa:2:30:10:8:true
TrimmomaticPE: Started with arguments:
 -threads 6 -trimlog ATACseq.txt -phred33 bulbul_R1.fastq.gz bulbul_R2.fastq.gz bulbul_R1_paired.fastq.gz bulbul_R1_unpaired.fastq.gz bulbul_R2_paired.fastq.gz bulbul_R2_unpaired.fastq.gz ILLUMINACLIP:/Users/gary/Applications/Trimmomatic-0.36/adapters/ATACseq.fa:2:30:10:8:true
Using Long Clipping Sequence: 'CAAGCAGAAGACGGCATACGAGATAGGAGTCCGTCTCGTGGGCTCGGAGATGT'
Using Long Clipping Sequence: 'AATGATACGGCGACCACCGAGATCTACACTCGTCGGCAGCGTCAGATGTG'
ILLUMINACLIP: Using 0 prefix pairs, 2 forward/reverse sequences, 0 forward only sequences, 0 reverse only sequences
Input Read Pairs: 26847985 Both Surviving: 26842135 (99.98%) Forward Only Surviving: 5290 (0.02%) Reverse Only Surviving: 557 (0.00%) Dropped: 3 (0.00%)
TrimmomaticPE: Completed successfully

Supplementary Table 1 of Buenrostro, et al (2013): Oligo designs. A list of ATAC-seq oligos used for PCR. Buenrostro JD, Giresi PG, Zaba LC, Chang HY, Greenleaf WJ. Transposition of native chromatin for fast and sensitive epigenomic profiling of open chromatin, DNA-binding proteins and nucleosome position. Nat Methods 2013;10(12):1213-8.

Ad1_noMX:
AATGATACGGCGACCACCGAGATCTACACTCGTCGGCAGCGTCAGATGTG

Ad2.1_TAAGGCGA
CAAGCAGAAGACGGCATACGAGATTCGCCTTAGTCTCGTGGGCTCGGAGATGT
Ad2.2_CGTACTAG
CAAGCAGAAGACGGCATACGAGATCTAGTACGGTCTCGTGGGCTCGGAGATGT
Ad2.3_AGGCAGAA
CAAGCAGAAGACGGCATACGAGATTTCTGCCTGTCTCGTGGGCTCGGAGATGT
Ad2.4_TCCTGAGC
CAAGCAGAAGACGGCATACGAGATGCTCAGGAGTCTCGTGGGCTCGGAGATGT
Ad2.5_GGACTCCT
CAAGCAGAAGACGGCATACGAGATAGGAGTCCGTCTCGTGGGCTCGGAGATGT
Ad2.6_TAGGCATG
CAAGCAGAAGACGGCATACGAGATCATGCCTAGTCTCGTGGGCTCGGAGATGT
Ad2.7_CTCTCTAC
CAAGCAGAAGACGGCATACGAGATGTAGAGAGGTCTCGTGGGCTCGGAGATGT
Ad2.8_CAGAGAGG
CAAGCAGAAGACGGCATACGAGATCCTCTCTGGTCTCGTGGGCTCGGAGATGT
Ad2.9_GCTACGCT
CAAGCAGAAGACGGCATACGAGATAGCGTAGCGTCTCGTGGGCTCGGAGATGT
Ad2.10_CGAGGCTG
CAAGCAGAAGACGGCATACGAGATCAGCCTCGGTCTCGTGGGCTCGGAGATGT
Ad2.11_AAGAGGCA
CAAGCAGAAGACGGCATACGAGATTGCCTCTTGTCTCGTGGGCTCGGAGATGT
Ad2.12_GTAGAGGA
CAAGCAGAAGACGGCATACGAGATTCCTCTACGTCTCGTGGGCTCGGAGATGT
Ad2.13_GTCGTGAT
CAAGCAGAAGACGGCATACGAGATATCACGACGTCTCGTGGGCTCGGAGATGT
Ad2.14_ACCACTGT
CAAGCAGAAGACGGCATACGAGATACAGTGGTGTCTCGTGGGCTCGGAGATGT
Ad2.15_TGGATCTG
CAAGCAGAAGACGGCATACGAGATCAGATCCAGTCTCGTGGGCTCGGAGATGT
Ad2.16_CCGTTTGT
CAAGCAGAAGACGGCATACGAGATACAAACGGGTCTCGTGGGCTCGGAGATGT
Ad2.17_TGCTGGGT
CAAGCAGAAGACGGCATACGAGATACCCAGCAGTCTCGTGGGCTCGGAGATGT
Ad2.18_GAGGGGTT
CAAGCAGAAGACGGCATACGAGATAACCCCTCGTCTCGTGGGCTCGGAGATGT
Ad2.19_AGGTTGGG
CAAGCAGAAGACGGCATACGAGATCCCAACCTGTCTCGTGGGCTCGGAGATGT
Ad2.20_GTGTGGTG
CAAGCAGAAGACGGCATACGAGATCACCACACGTCTCGTGGGCTCGGAGATGT
Ad2.21_TGGGTTTC
CAAGCAGAAGACGGCATACGAGATGAAACCCAGTCTCGTGGGCTCGGAGATGT
Ad2.22_TGGTCACA
CAAGCAGAAGACGGCATACGAGATTGTGACCAGTCTCGTGGGCTCGGAGATGT
Ad2.23_TTGACCCT
CAAGCAGAAGACGGCATACGAGATAGGGTCAAGTCTCGTGGGCTCGGAGATGT
Ad2.24_CCACTCCT
CAAGCAGAAGACGGCATACGAGATAGGAGTGGGTCTCGTGGGCTCGGAGATGT

Best,

Gary

[h.mon]

Please use the code button to format code, commands and such.

I formatted part of your post, to provide an example.

[Gary]

Many thanks for your information.

[ATpoint]

By the way, if you quote content from another threat, please use to " quote function in the formatting bar. I see you quoted a comment of mine but did not indicate it as such. Not a serious problem, but it helps people distinguish your content from those of other threads. In this case:

The only odd thing I see is that you trimmed for the TruSeq instead of the Nextera adapter.

[Gary]

Dear ATpoint, Sure, I have quoted them. Many thanks for your help. However, I don't understand that I trimmed the TruSeq instead of the Nextera adapter. Could you explain it more clearly to me? Thank you so much. Best, Gary

[ATpoint]

The only odd thing I see is that you trimmed for the TruSeq instead of the Nextera adapter.

is a quote from a previous thread which you did not indicate. It has nothing to do with this thread here.

CTGTCTCTTATACACATCT is for ATAC-seq

Answer 1

[ATpoint]

The transposase introduces the Nextera adapter sequence, therefore use CTGTCTCTTATACACATCT for both the forward and reverse trimming.

[liyc.stjude]

Is any TN5 requiring this sequence to be functional? CTGTCTCTTATACACATCT

Is there any paper about this sequence and how did people identify this sequence?

Thanks, Yichao