> ## Documentation Index
> Fetch the complete documentation index at: https://primerlab-genomic.mintlify.site/llms.txt
> Use this file to discover all available pages before exploring further.

# Sequence Handling

> Feature documentation: Sequence Handling

PrimerLab v0.1.6 introduces enhanced sequence handling for real-world genomic data.

## IUPAC Ambiguous Codes

IUPAC ambiguous nucleotide codes are automatically detected and handled:

| Code | Meaning    | Bases      |
| ---- | ---------- | ---------- |
| R    | puRine     | A or G     |
| Y    | pYrimidine | C or T     |
| W    | Weak       | A or T     |
| S    | Strong     | G or C     |
| K    | Keto       | G or T     |
| M    | aMino      | A or C     |
| B    | not A      | C, G, or T |
| D    | not C      | A, G, or T |
| H    | not G      | A, C, or T |
| V    | not T      | A, C, or G |

### Behavior

When IUPAC codes are detected:

1. **Warning displayed** to user
2. **Codes converted to N** (masked)
3. **Regions excluded** from primer placement

```bash theme={null}
$ primerlab stats sequence_with_iupac.fasta

📊 Sequence Statistics: TEST_IUPAC
=============================================
  Length:      500 bp
  GC Content:  48.20%
  ⚠️  IUPAC codes: ['R', 'Y'] (6 bp)
      → Will be converted to N during design
=============================================
```

## RNA Sequences

RNA sequences (containing Uracil) are automatically detected and converted:

| Detected      | Converted To       |
| ------------- | ------------------ |
| U (Uracil)    | T (Thymine)        |
| u (lowercase) | t (then uppercase) |

### Behavior

When RNA is detected:

1. **Warning displayed** to user
2. **U → T conversion** applied
3. **Design proceeds** with DNA sequence

```bash theme={null}
$ primerlab stats rna_sequence.fasta

📊 Sequence Statistics: TEST_RNA
=============================================
  Length:      300 bp
  ⚠️  RNA detected (contains U)
      → Will be converted to T during design
=============================================
```

## Sequence Validation

PrimerLab validates sequences before design:

| Check      | Requirement        | Error Code              |
| ---------- | ------------------ | ----------------------- |
| Length     | ≥ 50 bp            | ERR\_SEQ\_TOO\_SHORT    |
| Characters | A, T, G, C, N only | ERR\_SEQ\_INVALID\_CHAR |
| Non-empty  | Sequence provided  | ERR\_SEQ\_EMPTY         |

## Use `primerlab stats`

Always check your sequence before design:

```bash theme={null}
primerlab stats input.fasta
```