Reducing DNA damage during sample preparation and sequencing using siderophore chelators
Embodiments disclosed herein provide reagents and kits for nucleic acid preparation comprising a siderophore. Embodiments disclosed herein provide methods for preparing a nucleic acid library, which comprise: providing a plurality of nucleic acid molecules from a sample; and manipulating the plurality of nucleic acid molecules in a reagent for nucleic acid preparation comprising a siderophore. Further, embodiments disclosed herein provide methods for reducing oxidative damage to a nucleic acid molecule or increasing the Q (phred) score of a sequencing reaction, which methods comprise preparing the nucleic acid molecule in the absence of EDTA.
1. A composition for nucleic acid preparation comprising:
a reagent for nucleic acid preparation comprising a siderophore and a buffer, wherein the siderophore is present in the reagent at a concentration of 0.01 mM to 1 mM;
a plurality of oligonucleotides covalently bound to a solid support;
a plurality of nucleic acid molecules from a sample, wherein the plurality of nucleic acid molecules are hybridized to the plurality of oligonucleotides covalently bound to the solid support; and
an isolated enzyme selected from the group consisting of a DNA polymerase and a ligase;
wherein the composition does not contain EDTA, EGTA or DTPA.
2. The reagent of claim 1 , wherein the siderophore is a bacterial siderophore.
3. The reagent of claim 1 , wherein the siderophore is a desferrioxamine B (DFO-B).
4. The composition of claim 3 , wherein the DFO-B is present in the reagent at a concentration of 0.1 mM to 1 mM.
5. The composition of claim 3 , wherein the DFO-B is a mesylate salt.
6. The composition of claim 1 , wherein the enzyme is a ligase.
7. The composition of claim 1 , wherein the enzyme is a DNA polymerase.
8. The composition of claim 1 , wherein the siderophore is selected from the group consisting of desferrioxamine B, desferrioxamine E, fusarinine C, ornibactin, rhodotorulic acid, enterobactin, bacillibactin, vibriobactin, azotobactin, pyoverdine, yersiniabactin, mycobactin, ferrichrome, coprogen, arthrobactin, ferrichrome A, 2,3-dihydroxybenzoylserine, avenic acid, rhizoferrin, amonabactin T, and rhizoferrin.
9. The composition of claim 1 , wherein the solid support comprises a plurality of beads.
10. The composition of claim 1 , wherein the solid support comprises a plurality of magnetic beads.
11. The composition of claim 1 , wherein the sample comprises a biological sample.
12. The composition of claim 1 , wherein the sample is an artificial sample.