diff --git a/python/src/pyseqspec.rs b/python/src/pyseqspec.rs
index f0092bf..711b775 100644
--- a/python/src/pyseqspec.rs
+++ b/python/src/pyseqspec.rs
@@ -125,9 +125,13 @@ impl Assay {
         self.0.delete_read(read_id);
     }
 
-    fn validate(&self, modality: &str, out_dir: PathBuf) -> Result<()> {
+    #[pyo3(
+        signature = (modality, out_dir, *, tolerance=0.2),
+        text_signature = "($self, modality, out_dir, *, tolerance=None)",
+    )]
+    fn validate(&self, modality: &str, out_dir: PathBuf, tolerance: f64) -> Result<()> {
         let modality = Modality::from_str(modality)?;
-        self.0.validate(modality, out_dir)
+        self.0.validate(modality, out_dir, tolerance)
     }
 
     /*
diff --git a/seqspec/Cargo.toml b/seqspec/Cargo.toml
index 50d2e1d..90cbec9 100644
--- a/seqspec/Cargo.toml
+++ b/seqspec/Cargo.toml
@@ -12,7 +12,6 @@ log = "0.4"
 hamming = "0.1"
 noodles = { version = "0.80", features = ["core", "fastq", "async"] }
 reqwest = { version = "0.12", features = ["blocking"] }
-rayon = "1.10"
 serde = { version = "1.0", features = ["derive", "rc"] }
 serde_yaml = "0.9"
 zstd = { version = "0.13", features = ["zstdmt"] }
diff --git a/seqspec/src/lib.rs b/seqspec/src/lib.rs
index ce4610f..ad97846 100644
--- a/seqspec/src/lib.rs
+++ b/seqspec/src/lib.rs
@@ -3,7 +3,6 @@ mod region;
 pub mod utils;
 
 use log::warn;
-use rayon::iter::{IndexedParallelIterator, IntoParallelRefIterator, IntoParallelRefMutIterator, ParallelIterator};
 use read::{ReadSpan, RegionIndex, ValidateResult};
 pub use read::{Read, File, UrlType, Strand};
 use read::ReadValidator;
@@ -288,7 +287,7 @@ impl Assay {
 
     /// Validate reads in the sequence spec based on the fixed sequences and
     /// save the valid and invalid reads to different files.
-    pub fn validate<P: AsRef<Path>>(&self, modality: Modality, dir: P) -> Result<()> {
+    pub fn validate<P: AsRef<Path>>(&self, modality: Modality, dir: P, tolerance: f64) -> Result<()> {
         fs::create_dir_all(&dir)?;
         let (mut readers, reads): (Vec<_>, Vec<_>) = self.iter_reads(modality).flat_map(|read| {
             let reader = read.open()?;
@@ -307,6 +306,7 @@ impl Assay {
                 ReadValidator::new(region)
                     .with_range(range.start as usize ..range.end as usize)
                     .with_strand(read.strand)
+                    .with_tolerance(tolerance)
             }).collect::<Vec<_>>()
         ).collect();
 
@@ -335,7 +335,7 @@ impl Assay {
                 break;
             }
 
-            let valid = records.par_iter().zip(validators.par_iter_mut()).all(|(record, validators)|
+            let valid = records.iter().zip(validators.iter_mut()).all(|(record, validators)|
                 validators.iter_mut().all(|validator| {
                     match validator.validate(record.sequence()) {
                         ValidateResult::OnlistFail | ValidateResult::Valid => true,
diff --git a/seqspec/src/read.rs b/seqspec/src/read.rs
index a8ea143..51e2f00 100644
--- a/seqspec/src/read.rs
+++ b/seqspec/src/read.rs
@@ -288,7 +288,7 @@ pub(crate) struct ReadValidator<'a> {
     range: Option<Range<usize>>,
     n_total: usize,
     n_matched: usize,
-    onlist: Option<HashSet<String>>,
+    onlist: Option<HashSet<Vec<u8>>>,
     strand: Strand,
     tolerance: f64,
 }
@@ -329,6 +329,14 @@ impl<'a> ReadValidator<'a> {
         self
     }
 
+    pub fn with_tolerance(mut self, tolerance: f64) -> Self {
+        if tolerance < 0.0 || tolerance > 1.0 {
+            panic!("Tolerance must be between 0 and 1");
+        }
+        self.tolerance = tolerance;
+        self
+    }
+
     pub fn frac_matched(&self) -> f64 {
         self.n_matched as f64 / self.n_total as f64
     }
@@ -344,13 +352,18 @@ impl<'a> ReadValidator<'a> {
         } else {
             seq
         };
+        let seq = if self.strand == Strand::Neg {
+            crate::utils::rev_compl(seq)
+        } else {
+            seq.to_vec()
+        };
         let len = seq.len();
         if len < self.region.min_len as usize {
             ValidateResult::TooShort((seq.len(), self.region.min_len as usize))
         } else if seq.len() > self.region.max_len as usize {
             ValidateResult::TooLong((seq.len(), self.region.max_len as usize))
         } else if self.sequence_type() == SequenceType::Fixed {
-            let d = hamming::distance(seq, self.region.sequence.as_bytes()) as f64;
+            let d = hamming::distance(&seq, self.region.sequence.as_bytes()) as f64;
             if d <= self.tolerance * len as f64 {
                 self.n_matched += 1;
                 ValidateResult::Valid
@@ -358,24 +371,11 @@ impl<'a> ReadValidator<'a> {
                 ValidateResult::MisMatch(d as usize)
             }
         } else if let Some(onlist) = &self.onlist {
-            match self.strand {
-                Strand::Neg => {
-                    let seq = crate::utils::rev_compl(seq);
-                    if onlist.contains(std::str::from_utf8(&seq).unwrap()) {
-                        self.n_matched += 1;
-                        ValidateResult::Valid
-                    } else {
-                        ValidateResult::OnlistFail
-                    }
-                },
-                Strand::Pos => {
-                    if onlist.contains(std::str::from_utf8(seq).unwrap()) {
-                        self.n_matched += 1;
-                        ValidateResult::Valid
-                    } else {
-                        ValidateResult::OnlistFail
-                    }
-                },
+            if onlist.contains(&seq) {
+                self.n_matched += 1;
+                ValidateResult::Valid
+            } else {
+                ValidateResult::OnlistFail
             }
         } else {
             self.n_matched += 1;
diff --git a/seqspec/src/region.rs b/seqspec/src/region.rs
index c803c4b..efb7f60 100644
--- a/seqspec/src/region.rs
+++ b/seqspec/src/region.rs
@@ -256,11 +256,11 @@ pub struct Onlist {
 }
 
 impl Onlist {
-    pub fn read(&self) -> Result<HashSet<String>> {
+    pub fn read(&self) -> Result<HashSet<Vec<u8>>> {
         let cache = Cache::new()?;
         let file = cache.cached_path(&self.url)?;
         let reader = std::io::BufReader::new(crate::utils::open_file_for_read(file));
-        Ok(reader.lines().map(|x| x.unwrap()).collect())
+        Ok(reader.lines().map(|x| x.unwrap().into_bytes()).collect())
     }
 
     pub(crate) fn normalize_path<P: AsRef<Path>>(&mut self, work_dir: P) -> Result<()> {