Sonication Tests for Sea Star Library Prep

Testing Sonication Protocol for 350-500bp Range for Sea Star DNA

20210618 2 minutes and 2.5 minutes at 25% amplitude

Samples for 2 minutes:

Sample	ul DNA for 500ng	ul 10mM Tris HCl to 80ul
CHSB 002	10.99	69.01
CPAB2 001	14.71	65.29
CPDB 002	8.12	71.88
CPBP 004	8.91	71.09

• Thaw DNA on ice
• Set up sonicator (see protocol)
• Set program to 2 minutes with 25% amplitude and 15 second on and 15 seconds off pulse
• Aliquot 10uM tris HCl into sonicator tubes as in table above
• Vortex and spin down DNA
• Add DNA into sonciator tubes as in table above
• Vortex and spin down sonicator tubes
• Place in sonicator and start program
• After 1 minute pause program, take tubes out and spin down, put back in, and continue program
• Spin down tubes and keep on ice when done

Samples for 2 minutes 30 seconds:

Sample	ul DNA for 500ng	ul 10mM Tris HCl to 80ul
CPOI 004	9.63	70.37
CPBP 009	10.85	69.15
CPAB 005	7.2	72.8
CPAB2 008	12.56	67.44

• Thaw DNA on ice
• Set program to 2 minutes 30 seconds with 25% amplitude and 15 second on and 15 seconds off pulse
• Aliquot 10uM tris HCl into sonicator tubes as in table above
• Vortex and spin down DNA
• Add DNA into sonciator tubes as in table above
• Vortex and spin down sonicator tubes
• Place in sonicator and start program
• After 1 minute 15 seconds: pause program, take tubes out and spin down, put back in, and continue program
• Spin down tubes and keep on ice when done

1X Bead Cleanup following protocol: samples we resuspended in 12.5ul of 10mM tris HCl.

1% Gel following protocol: gel was ran for 2 hours at 80V.

20210621 1 minute and 1.5 minutes at 35% amplitude

Samples for 1 minute:

Sample	ul DNA for 500ng	ul 10mM Tris HCl to 80ul
CHSB 004	12.2	67.8
CHSY 004	12.47	67.53
CPAB 003	8.08	71.92
CPBP 010	9.78	70.22

• Thaw DNA on ice
• Set up sonicator (see protocol)
• Set program to 1 minute with 35% amplitude and 15 second on and 15 seconds off pulse
• Aliquot 10uM tris HCl into sonicator tubes as in table above
• Vortex and spin down DNA
• Add DNA into sonciator tubes as in table above
• Vortex and spin down sonicator tubes
• Place in sonicator and start program
• After 30 seconds pause program, take tubes out and spin down, put back in, and continue program
• Spin down tubes and keep on ice when done

Samples for 1 minute 30 seconds:

Sample	ul DNA for 500ng	ul 10mM Tris HCl to 80ul
CPDB 009	10.31	69.69
CPBO 006	9.75	70.25
CPBP 007	13.55	66.45
CPOI 002	11.9	68.1

• Thaw DNA on ice
• Set program to 1 minute 30 seconds with 35% amplitude and 15 second on and 15 seconds off pulse
• Aliquot 10uM tris HCl into sonicator tubes as in table above
• Vortex and spin down DNA
• Add DNA into sonciator tubes as in table above
• Vortex and spin down sonicator tubes
• Place in sonicator and start program
• After 45 seconds: pause program, take tubes out and spin down, put back in, and continue program
• Spin down tubes and keep on ice when done

1X Bead Cleanup following protocol: samples we resuspended in 12.5ul of 10mM tris HCl.

1% Gel following protocol: gel was ran for 2 hours at 80V.

Written on June 21, 2021