Acoustic energy mediation of genetic fragmentation
Method and apparatus for controlling acoustic treatment of a sample to mediate a tagmentation process used on double stranded DNA.
1. A method for processing a sample containing genomic material, comprising:
providing a sample in a vessel, the sample including double stranded DNA material with segments having a base pair length in excess of a starting base pair length;
subjecting the sample to acoustic energy to cause random shearing of the segments of double stranded DNA material in the sample to form fragments of the double stranded DNA material having a base pair length of a final base pair length, the starting base pair length being two or more times the final base pair length; and
providing a hyperactive mutant of Tn5 transposase and oligonucleotide material with the fragments of double stranded DNA material to cut the fragments of double stranded DNA material with the Tn5 transposase and insert oligonucleotide material into the fragments of double stranded DNA material at areas cut by the Tn5 transposase.
2. The method of claim 1 , wherein the step of providing a hyperactive mutant of Tn5 transposase and oligonucleotide material with the fragments of double stranded DNA material occurs before the step of subjecting the sample to acoustic energy.
3. The method of claim 1 , further comprising:
subjecting the sample to acoustic energy to mix the hyperactive mutant of Tn5 transposase, oligonucleotide material, and the fragments of double stranded DNA material after the step of subjecting the sample to acoustic energy to cause shearing.
4. The method of claim 1 , wherein the oligonucleotide material includes synthetic oligonucleotides.
5. The method of claim 1 , wherein the sample volume is about 15 microliters.
6. The method of claim 1 , wherein the vessel volume is about 100 microliters.
7. The method of claim 1 , wherein the starting base pair length is in excess of 10000 bp.
8. The method of claim 1 , wherein the final base pair length is less than 3000 bp.
9. The method of claim 1 , wherein the final base pair length is between 1000 bp and 1500 bp.
10. The method of claim 1 , wherein the step of subjecting the sample to acoustic energy reduces a viscosity of the sample so as to enhance enzyme interaction that occurs during the step of providing a hyperactive mutant of Tn5 transposase and oligonucleotide material.
11. The method of claim 1 , wherein the subjecting step is performed over a time period of 30-200 seconds.
12. The method of claim 1 , wherein the step of providing a hyperactive mutant of Tn5 transposase and oligonucleotide material with the fragments of double stranded DNA material occurs after the step of subjecting the sample to acoustic energy.
13. The method of claim 12 , further comprising:
subjecting the sample to acoustic energy to mix the hyperactive mutant of Tn5 transposase, oligonucleotide material, and the fragments of double stranded DNA material after the step of subjecting the sample to acoustic energy to cause shearing.
14. The method of claim 1 , wherein the oligonucleotide material includes synthetic oligonucleotides.
15. A method for processing a sample containing genomic material, comprising:
providing a sample in a vessel, the sample including double stranded DNA material with segments having a base pair length in excess of 3000 bp;
subjecting the sample to acoustic energy to cause random shearing of the segments of double stranded DNA material in the sample to form fragments of the double stranded DNA material having a base pair length less than 3000 bp; and
providing a hyperactive mutant of Tn5 transposase and oligonucleotide material with the fragments of double stranded DNA material to cut the fragments of double stranded DNA material with the Tn5 transposase and insert oligonucleotide material into the fragments of double stranded DNA material at areas cut by the Tn5 transposase, wherein a concentration of the Tn5 transposase is less than that needed to perform a tagmentation process using techniques without acoustic energy treatment.
16. A method for processing a sample containing genomic material, comprising:
providing a sample in a vessel, the sample including double stranded DNA material with segments having a base pair length in excess of 3000 bp;
subjecting the sample to acoustic energy to cause random shearing of the segments of double stranded DNA material in the sample to form fragments of the double stranded DNA material having a base pair length less than 3000 bp; and
providing a hyperactive mutant of Tn5 transposase and oligonucleotide material with the fragments of double stranded DNA material to cut the fragments of double stranded DNA material with the Tn5 transposase and insert oligonucleotide material into the fragments of double stranded DNA material at areas cut by the Tn5 transposase, wherein the fragments of double stranded DNA material having inserted oligonucleotide material have a tighter size distribution than that resulting from standard tagmentation processes performed in the absence of acoustic treatment.