BioCurious: DIY Ultrasonic Transformation

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(Materials)
(Procedure)
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## 72 °C 5 min
## 72 °C 5 min
## 12 °C hold
## 12 °C hold
 +
 +
 +
----
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Still in progress, above kept for template.
 +
 +
 +
Sonicate each tube for 10 secs at only available setting on Sonicator (on/off)except 2nd control tube with plasmid and Al
 +
 +
#Test transform Tube 1 with Al placed in middle of Transducer bump, flat glass  bottom down
 +
#Test transform Tube 2 without Al placed in middle of Transducer bump same way
 +
#Test transform Tube 3 with Al placed in a side of the bath away from transducer
 +
#Test transform Tube 4 without Al placed in a side of the bath away from transducer
 +
#Control Tube 1 without plasmid and Al placed in middle of transducer bump
 +
#Control Tube 2 with plasmid and Al left alone
 +
 +
Post sonication
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#Add 1 mL LB solution
 +
#Incubate at 37C 1 hr on a bath-shaker
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        ##Plate LB/AmpC after 1 hour and incubate for a day
 +
        ##Check plates for colony in UV light next day
==Notes==
==Notes==

Revision as of 22:33, 18 March 2013

Contents

Overview

Experiment: Transform HB101 Strain with pGlo plasmid using ultrasound device

Materials

For an XX μL transformation:

Equipment:

Off-the-shelf Ultrasonic Jewellery cleaning box with ~ 1L bath Strains DNA reagent in Dark Glass flat bottom tubes 5 mL with Plastic Caps

Sonicator Essentials:

42 KHz 35 watt peak 20 sq cm transducer area in center of bath Bath filled with tap water at room temperature

Procedure

  1. In a PCR tube, mix the components on ice in the order they are listed above.
  2. Perform thermocycling program
    1. 95 °C 5 min
    2. 95 °C 30 s
    3. TH 30 s
    4. 72 °C 1 min for each 1 kb PCR product
    5. Repeat steps 2-4 a total of 12-36 times (24 is standard).
    6. 72 °C 5 min
    7. 12 °C hold



Still in progress, above kept for template.


Sonicate each tube for 10 secs at only available setting on Sonicator (on/off)except 2nd control tube with plasmid and Al

  1. Test transform Tube 1 with Al placed in middle of Transducer bump, flat glass bottom down
  2. Test transform Tube 2 without Al placed in middle of Transducer bump same way
  3. Test transform Tube 3 with Al placed in a side of the bath away from transducer
  4. Test transform Tube 4 without Al placed in a side of the bath away from transducer
  5. Control Tube 1 without plasmid and Al placed in middle of transducer bump
  6. Control Tube 2 with plasmid and Al left alone

Post sonication

  1. Add 1 mL LB solution
  2. Incubate at 37C 1 hr on a bath-shaker
        ##Plate LB/AmpC after 1 hour and incubate for a day
        ##Check plates for colony in UV light next day

Notes

Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!

  1. List troubleshooting tips here.
  2. You can also link to FAQs/tips provided by other sources such as the manufacturer or other websites.
  3. Anecdotal observations that might be of use to others can also be posted here.

Please sign your name to your note by adding '''*~~~~''': to the beginning of your tip.

References

Relevant papers and books

  1. Goldbeter, A and Koshland, DE (1981) - Proc Natl Acad Sci U S A 78(11) 6840-4 PMID 6947258
  2. Jacob, F and Monod, J J (1961) - Mol Biol 3(3) 318-56 PMID 13718526
  3. Ptashne, M (2004) Genetic Switch: Phage Lambda Revisited - Cold Spring Harbor Laboratory Press ISBN 0879697164

Contact

  • Who has experience with this protocol?

or instead, discuss this protocol.

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