DTT (Dithiothreitol)
Definition
A reducing agent that cleaves the disulphide bonds in the sperm protamine matrix, allowing proteinase K and SDS to lyse the sperm nuclei in the second step of differential extraction.
- Chemical role
- Reducing agent
- Target
- Disulphide bonds in sperm protamine matrix
- Used in
- Second step of differential extraction
- Works alongside
- Proteinase K and SDS
Common questions
Why can't proteinase K alone lyse sperm cells in a sexual assault sample?+
Sperm nuclei are unusually resistant to standard lysis because their DNA is packaged with protamines cross-linked by disulphide bonds, a structure not present in typical somatic cells. Proteinase K and SDS efficiently lyse epithelial cells but cannot break those disulphide bonds without DTT, which is why differential extraction uses DTT specifically at the sperm-lysis stage.
What happens if the DTT step is skipped or incomplete in differential extraction?+
Incomplete disulphide reduction leaves sperm cells partially or wholly unlysed, so their DNA is under-recovered in the sperm fraction, which can produce a weak or missing male profile and an inaccurate picture of the mixture even when sperm cells are visibly present on the slide.
Related terms
- Differential Extraction
- A two-step DNA extraction procedure used when a stain contains both sperm cells and non-sperm epithelial cells. The first step lyses epithelial...
- Erase Sperm Isolation Kit
- An immunomagnetic sperm-cell isolation system using paramagnetic beads coated with antibodies against sperm-specific surface antigens. Achieves greater than 99% sperm recovery with...
- Female Fraction
- The lysate produced by the first (no-DTT) lysis step of differential extraction. Contains DNA from epithelial cells and other non-sperm cellular material...
- Laser Capture Microdissection (LCM)
- A UV-laser instrument technique for physically isolating individual cells or clusters of cells from a stained slide into a capture tube, without...
- Male Fraction (Sperm Fraction)
- The lysate produced by the second (DTT + proteinase K) lysis step after washing the sperm pellet. Ideally contains predominantly the suspect's...
- Protamine
- Small arginine-rich proteins that replace histones in the nuclei of spermatozoa during spermatogenesis, producing a hyper-compacted chromatin structure approximately six times denser...