Team:ZJU-China/Notebook/September

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Labnote

Biobrick group | Biofilm group

This group...........

May&June

In the two months we discussed some new ideas and finally decided our project
>> Click to see our brainstorm

July

In this month we......
>>Click to see

August

In this month we......
>>Click to see

September

In this month we......
>>Click to see

Week9

Day

Note

Aug.29th Monday

• Meeting. Summarize all the experiment work so far.

• Cut: 22M(X+P), 1C3(E+P)

Aug.30th Tuesday

Aug.31st Wednesday

• Cut vgb+YFP+tetR, fdhF+RFP+tetR.
• Run the digestion results

• Culture the positive results

Sept.1st Thursday

• Miniprep: Y#4, R#2/3/4/5
• Cut Y#4, R#2/3/4/5

•Run the gel.
• Send R#3,Y#4 sample to sequencing

• Hypoxia culture: RFP3, RFP4
• Preserve Y#4, R#2/3/4/5

Sept.2nd Friday

• Phusion PCR

• Hypoxia Culture: R, Y
• Run the PCR results. Bands are confirmed right.

• Purification: vgb, YFP, tetR.

Sept.3rd Saturday

• Culture: YFP, RFP, A25
• Culture in micro-oxygen: YFP, RFP, A25

• Culture in slope medium: RFP, YFP, A25
• Hypoxia culture: RFP, YFP, A25

• Check the fluorescence.
• Neither RFP in hypoxia or oxygen condition. Little YFP both in hypoxia and oxygen.

Sept.4th Sunday

• Miniprep: RFP3, YFP4

• Cut: RFP3, YFP4, !C3.

Week10

Day

Note

Sept.5th Monday

• Run the gel: Yu’s PCR results, 1C3, RFP3, YFP4
• Purification: RFP3, YFP4

• Ligation: RFP3+1C3, YFP4+1C3
• Hypoxia culture: RFP3, YFP4, A25.

• Transform the ligation results.
• Confirmed RFP expression only in hypoxia condition.

Sept.6th Tuesday

• Check the RFP, YFP plates.

• Colony PCR

Sept.7th Wednesday

• Culture YFP1/2, RFP3/4

Sept.8th Thurday

• Preserve the strain of RFP1, YFP1/2 in 1C3.
• Miniprep the remains.

• Cut: YFP1/2, RFP1

• Run the digestion results.
• Culture RFP2

Sept.9th Friday

• Miniprep: RFP2, YFP2, RFP1

• Cut them and run the gel.

Sept.10th Saturday

• Cut YFP(1A3), RFP(1A3) with S+E

• Purification: RFP(1A3), YFP(1A3)

• Ligation: RFP+1C3, YFP+1C3

• Transform the ligation results.

Sept.11th Sunday

• Check the plates.

• Culture lig RFY+YFP+IC3
• Culture 1K3

• Colony PCR: RFP(1C3)， YFP(1C3)

Week11

Day

Note

Sept.12th Monday

• Colony PCR: 1A3, 1K3, lig RFP+YFP+1C3.

• Run the PCR results.

• Miniprep: lig RFP+YFP+1C3

Sept.13th Tuesday

• Transform 2M, 2I, 6I, 23L.

• Purification: 1K3

Sept.14th Wednesday

• Miniprep: 20H, 20J, 22B

• Cut: 1K3 with E+P, lig3 with S+E, 12I(CFP) with P+X,

• Gel excision and /or purification: 1K3, lig3, 12I(CFP)

• Run the gel.
• Culture: 1I, 1K, 3L, 5E, 7C

• Miniprep: RFP+YFP+1C3.
• Run the gel of 1K3, lig3, CFP again.

• Gel excision and purification
• Ligation: Lig3+CFP+1K3

Sept.15th Thursday

• Preserve: 1I, 1K, 3C, 5E, 7C
• Transform the ligation results

• Colony PCR: 2M, 2I，6I，23L
• Cut: 20H, 20J, 22B with X+P

• Miniprep: 2M-1, 2I-1, 6I-1, 23L-1
• Cut: 2M, 2I with E+S

Sept.16th Friday

• Colony PCR: RFP+YFP+1K3

• Cut 13K with E+S, 10I with P+X, 1C3 with E+P

• Run the gel: 13K, 10I, 1C3, 2I, 2M, 20J, 22H, 22B

• Ligation: 13K+10I+1C3
• Culture: YFP(Amp), CFP(Amp)

Sept.17th Saturday

• Order primers for 20H, 20J
• Culture 20H-1, 20J-1, 22B-1

• Plate the ligation results
• Culture RFP+YFP+CFP+1K3

• Colony PCR: 3A, 1C3
• Preserve: 1K3, RFP+YFP+1C3.

Sept.18th Sunday

• Miniprep: 20J, 20H, RFP+YFP+CFP+1K3 1A3

• Run the gel of PCR results

• Cut RFP+YFP+CFP+1K3 with E+P
• Colony PCR: 13K+10I+1C3

• PCR: fdhF+rush+Vgb+tetR
• Preserve: 20H, 20J, 22B

• Run the gel: 20H, 20J, 22B
• Cut the bands and purification.

• Run the gel: 13K+10I+1C3, RFP+YFP+CFP+1K3
• Culture: 13K+10I+1C3
• Grad PCR: fdhF.

Week12

Day

Note

Sept.19th Monday

• Miniprep: 13K+10I+1C3, 1C3
• Run the gel of digestion results.

• Cut: 13K+10I+1C3, 1A3, fdhF

• Run the gel: 20H, 20J, rush+vgb, RYC+1K3, fdhF, 13K+10I, 1A3

• Cut and purification: fdhF, 13K+10I, 1A3
• Culture: CFP, YFP

• Purification: Rush PCR
• Cut: 22B with X, rush PCR results with S

Sept.20th Tuesday

• Run the 20H, 20J from PCR results.
• Transform: fdhF+13K+10I+1A3

• Purification: 20B, Rush
• Ligation: 22B+Rush

• Purification: ligation results
• Cut: the ligation results with X+S

• PCR 20H
• Self-ligation: rush+22M

Sept.21st Wednesday

• Colony PCR: fdhF+13K+10I+1A3 on plates

• Run the gel of PCR results

• Culture the positive results.

Sept.22nd Thursday

• Miniprep: fdhF+13K+10I+1A3
• Preserve the above.

• Cut and run the results: fdhF+13K+10I+1A3, 13K+10I+1C3

• Cut: RFP with E+S, YFP with X+P, 1K3 with E+P, 1C3 with E+P

• Purification the digestion results.

Sept.23rd Friday

• Ligate: RFP+1A3/ YFP+1C3/1K3, RFP+1A3/1C3

• Transform the ligation results.

Sept. 24th Saturday

• Order primers for 20H, 20J
• Culture 20H-1, 20J-1, 22B-1

• Plate the ligation results
• Culture RFP+YFP+CFP+1K3

• Colony PCR: 3A, 1C3
• Preserve: 1K3, RFP+YFP+1C3.

Sept.25th Sunday

• Miniprep: 20J, 20H, RFP+YFP+CFP+1K3 1A3

• Run the gel of PCR results

• Cut RFP+YFP+CFP+1K3 with E+P
• Colony PCR: 13K+10I+1C3

• PCR: fdhF+rush+Vgb+tetR
• Preserve: 20H, 20J, 22B

• Run the gel: 20H, 20J, 22B
• Cut the bands and purification.

• Run the gel: 13K+10I+1C3, RFP+YFP+CFP+1K3
• Culture: 13K+10I+1C3
• Grad PCR: fdhF.