Team:Paris Saclay/Notebook/August/19
Contents
Wednesday 19th August
Lab Work
Electrophoresis
by Pauline
Agarose gel 1%, Migration 90V
- BBa_K1707000 #2, #3, #4
- BBa_K1707013 #1
- BBa_K1707030 #1
- BBa_K1707019 #1
- BBa_K1707020 #2
- BBa_K1707021 #2
- BBa_K1707035 #1
- BBa_K1707036 #1
- BBa_K1707027 #1
- pIJ773
- BBa_K1707022 #1 and #2
- BBa_K1707023 #1 and #2
- BBa_K1707034 #1 and #2
Verification of PCR products, from left to right: 1. DNA Ladder, 2. BBa_K115017, 3. BBa_K1707039, 4. BBa_K1707000#2, 5. BBa_K1707000#3, 6. BBa_K1707000#4, 7. BBa_K1707013#1, 8. BBa_K1707030#1, 9. BBa_K1707019#1, 10. BBa_K1707020#2, 11. BBa_K1707021#2, 12. BBa_K1707027#1
Wells 1-2: Verification of PCR products, Well 3: Quantification, Wells 5-10: Verification by digestion with XbaI and PstI; from left to right: 1. BBa_K1707035#1, 2. BBa_K1707036#1, 3. BBa_PIJ773 (10%), 4. DNA Ladder, 5. BBa_K1707022#1, 6. BBa_K1707022#2, 7. BBa_K1707023#1, 8. BBa_K1707023#2, 9. BBa_K1707034#1, 10. BBa_K1707034#2
Digestion
by Pauline
- BBa_K1707022 #1 and BBa_K1707028 #1
- 10 µL plasmid
- 1 µL XbaI
- 1 µL PstI
- 2 µL Buffer FastDigest 10x
- 6 µL H2O
- BBa_K1707034 #1
- 10 µL plasmid
- 1 µL SpeI
- 2 µL PstI
- 2 µL Tango Buffer 10x
- 5 µL H2O
- BBa_K1707023 #1 and #2
- 5 µL plasmid
- 0,5 µL XbaI
- 0,5 µL PstI
- 1 µL Buffer FastDigest 10x
- 3 µL H2O
Incubation 37°C, 1h30
Electrophoresis
by Pauline
Biobricks:
- BBa_K1707023 #1 and #2
- BBa_K1707022 #1
- BBa_K1707028 #1
Agarose gel 1%, 90V
Wells 1-2: Verification by digestion with XbaI and PstI, Wells 4-5: Verification of gel purification; from left to right: 1. BBa_K1707023#1, BBa_K1707023#2, 3. DNA Ladder, 4. BBa_K1707022#1, 5. BBa_K1707028#1, 6. Empty, 7. Empty, 8. Empty, 9. Empty, 10. Empty
We can conclude that BBa_K1707023 #1 and #2 are OK. We can cut BBa_K170708 #1 and BBa_K1707022 #1 bands.PCR
by Pauline
Biobricks:
- BBa_K1707031 #7 to #26
We put 30 µL H2O in eqch tub. We add then the plasmid. And after, the mix.
Mix for each tub:
- 10 µL Buffer
- 0,25 µL Forward primer (iPS43)
- 0,25 µL Reverse primer (iPS44)
- 1 µL dNTP
- 0,25 µL GoTAQ
- 4 µL MgCl2
- 4,25 µL H2O
Digestion
by Pauline and Audrey
Biobricks: BBa_K1707004 #2
Mix:
- 15 µL plasmid
- 3 µL XbaI
- 1,5 µL SpeI
- 3 µL Tango Buffer 10x
- 7,5 µL H2O
Biobrick: BBa_K1707023 #1
Mix:
- 20 µL plasmid
- 2 µL PstI
- 1 µL SpeI
- 3 µL Tango Buffer 10x
- 4 µL H2O
Incubation 37°C, ON
Member present:
- Instructors: Claire
- Students: Pauline and Audrey
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t@iGEMParisSaclay
aUniversité Paris Sud
a91405 Orsay, France
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