Team:Paris Saclay/Notebook/August/20

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Thursday 20th August

Lab Work

Digestion for check

by Audrey

• BBa_K115017 (control +)
• BBa_J23101 + GFP (control -)
• BBa_K1707000 #2, #3 and #4
• BBa_K1707013 #1
• BBa_K1707030 #1
• BBa_K1707019 #1
• BBa_K1707020 #2
• BBa_K1707021 #2
• BBa_K1707027 #1
• BBa_K1707035 #1
• BBa_K1707036 #1

Mix for each tub:

• • 2 µL plasmid
  • 1 µL SpeI (Biolabs)
  • 1 µL BspE1 (Biolabs)
  • 2 µL Buffer 3 10x (Biolabs)
  • 5 µL H2O

Electrophoresis to check PCR products

by Audrey

Agarose gel 1%, Migration 90V

Biobricks:

• BBa_K1707031 #7 to #26

Verification of BBa_K1707031 PCR products, from left to right: 1. DNA Ladder, 2. #7, 3. #8, 4. #9, 5. #10, 6. #11, 7. #12, 8, #13, 9. #14, 10. #15, 11. #16, 12. #17

Verification of BBa_K1707031 PCR products, from left to right: 1. DNA Ladder, 2. #18, 3. #19, 4. #20, 5. #21, 6. #22, 7. #23, 8, #24, 9. #25, 10. #26

We can conclude that all clones aren't OK

Purification

by Pauline

• BBa_K1707022 #1
• BBa_K1707023 #1
• BBa_K1707004 #2
• BBa_K1707034 #1
• BBa_K1707028 #1

With Macherey Nagel purification kit

Electrophoresis

by Pauline

Digestion products:

• BBa_K115017 (control +)
• BBa_J23101 + GFP (control -)
• BBa_K1707000 #2, #3 and #4
• BBa_K1707013 #1
• BBa_K1707030 #1
• BBa_K1707019 #1
• BBa_K1707020 #2
• BBa_K1707021 #2
• BBa_K1707027 #1
• BBa_K1707035 #1
• BBa_K1707036 #1

Verification by digestion with SfoI and BspEI, 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 by digestion with SfoI and BspEI, Wells 4-8: Quantification; from left to right: 1. BBa_K1707035#1, 2. BBa_K1707036#1, 3. DNA Ladder, 4. BBa_K1707004#2, 5. BBa_K1707022#1, 6. BBa_K1707023#1, 7. BBa_K1707023#1, 8. BBa_K1707034#1, 9. Empty, 10. Empty

Quantification:

• BBa_K1707022 #1: ?
• BBa_K1707023 #1: nothing
• BBa_K1707004 #2: nothing
• BBa_K1707034 #1: 20 µg/µL
• BBa_K1707028 #1: 10 µg/µL

New culture

by Pauline

• BBa_K1707004 #2

In 5mL LB + 5µL Ampicillin

Ligation

by Pauline

• BBa_K1707037 = BBa_K1707028 + BBa_K1707034

Mix:

• 8 µL BBa_K1707028
• 10 µL BBa_K1707034
• 2 µL Ligase
• 2,5 µL Buffer
• 2,5 µL H2O

Incubation 4°C, ON

Quantification

by Pauline

BIobricks:

• BBa_K1707022 #1
• BBa_K1707023 #1

Quantification, from left to right: 1. DNA Ladder, 2. BBa_K1707022#1, 3. BBa_K1707023#1, 4. Empty, 5. Empty, 6. Empty, 7. Empty, 8. Empty, 9. Empty, 10. Empty, 11. Empty, 12. Empty

We can't see anything

Member present:

• Instructors: Claire
• Students: Pauline and Audrey

Back to the calendar

Contact
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t@iGEMParisSaclay
aUniversité Paris Sud
a91405 Orsay, France Copyright © 2015 iGEM Paris Saclay. All rights reserved All inspirations encouraged.
*This wiki was conceived during long nights of work and a great coffee experience in the beautiful Orsay, France.