Team:OUC-China/Lab note

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Revision as of 08:29, 26 September 2013

Lab-note

June 26th

Person: Wenjun Wang

Experiment:
1. Get the AMB-1 bacteria strain from Nanjing University.
2. Keep the strain in 30℃.

June 30th

Person: Wengjun Wang 　　　

Experiment:
1. PCR: Get 16srDNA sequence from AMB-1 bacteria genome to detect the strain.
2. Agarose electrophoresis: detect the PCR product.
3. Transform: transform the ligastion product into Top 10 to prepare for sequence. 　　　

July 13th ~ 15th

Person: Wenjun Wang

Experiment:
1. Get the AMB-1 bacteria strain from Nanjing University.
2. Keep the strain in 30℃.

June 30th

Person: Kaili Qin 　　　

Experiment:
　　1. Equip the medium: Equip the medium of Magnetospirillum Magneticum AMB-1.
　　　

July 20th

Person: Wenjun Wang 　　　

Experiment:
1. PCR: Get mamJ sequence from PCR product to finish the Over-Lap PCR.
2. Agarose electrophoresis: detect the plasmid PCR product.
3. Transform: transform the ligastion product into Top 10 to prepare for sequence.
4. PCR: Get mamK sequence from PCR product to finish the Over-Lap PCR.
5. Agarose electrophoresis: detect the PCR product.
6. Product purification: purify the PCR product.
7. PCR: Get mamAB sequence from AMB-1 bacteria genome. 　　　

Person: Qianyun Lu 　　　

1. Culture the E.coli cells: Psb1c3.
　　　

July 21st

Person: Wenjun Wang 　　　

Experiment:
1. Agarose electrophoresis: detect the plasmid PCR product.
2. PCR: Get mamB sequence from PCR product.
3. PCR: Get mamQ sequence from PCR product.
4. PCR: Get mamJ sequence from PCR product to finish the Over-Lap PCR.
5. Agarose electrophoresis: detect the plasmid PCR product.
6. Product purification: purify the PCR product. 　　　

Person: Yu Wang 　　　

Experiment:
1. Bacterination: E.coli with part K1059003-B0035-GFPlva-K1059004-B0015 pSB1AK3 as backbone; E.coli with RFP, pSB1C3 as backbone. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Plasmid miniprep: plasmid Extraction of PSB1C3 for 5 tubes.
2. Electrophoresis: gel is good.
3. Equip the medium: Equip the medium of LB (solid). 　　　

Person:Qianyun Lu 　　　

Experiment:
1. PCR production purification: 16SrRNA from AMB-1.
2. SDS page gel electrophoresis: the purified PCR production. 　　　

Person:Xiaodong Zhong 　　　

Experiment:
1. PCR genome for mamI gene & PCR purification.
2. PCR genome for mamB gene. 　　　

July 22nd

Person:Xue Sun 　　　

Experiment:
1. Plasmid miniprep: Plasmid pSB1AK3 with K1059003-B0035-GFPlva-K1059004-B0015, and pSB1C3 with RFP.
2. Enzyme digestion.
3. Gel extraction.
4. Recombination: Get part K1059003-B0035-GFPlva-K1059004-B0015, pSb1C3 as backbone. 　　　

Person:Kaili Qin 　　　

Experiment:
1.Enzyme digestion: enzyme digestion of PSB1C3 for 25ul system.
2.Gel recovery: gel recovery of the product of enzyme digestion.
3.Bacterination: bacterinate PSB1C3-RFP, RNA1 into tubes of 10ml.
4.Pick bacteria: pick bacteria of PSB1C3-RFP for conservation.
5.Genomic DNA extraction: extracting genomic DNA of E.coli.
6. Electrophoresis: gel is bad.
7. Bacterination: bacterinate PSB1C3-RFP, RNA1 into tubes of 10ml for 4 tubes. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Ligation: ligate the purified PCR production with T vector.
2. Transformation of Top10. 　　　

July 23rd

Person:Qianyun Lu 　　　

Experiment:
1. Extraction of genomic DNA from E.coli.
2. Pick the bacteria and culture the E.coli cells for four hours.
3. PCR: the cultured E.coli cells.
4. SDS page gel electrophoresis: PCR production. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Plasmid miniprep: plasmid Extraction of PSB1C3 -RFP for 5 tubes.
2. Enzyme digestion: enzyme digestion of PSB1C3 for 50ul system of 3 tubes, use EcoR1, Pst1.3. Electrophoresis: gel is good.
4. Genomic DNA extraction: extracting genomic DNA of E.coli.
5. Electrophoresis: gel is bad.
6. Gel recovery: gel recovery of PSB1C3.
7. Electrophoresis: gel is good for 50ng/ul each.
8. Bacterination: bacterination of E.coli for 6 tubes. 　　　

Person:Qiu Wang 　　　

Experiment:
1. Transformation: Plate cultivation for 14h. 　　　

July 24th

Person:Qianyun Lu 　　　

Experiment:
1. Extraction of genomic DNA from E.coli.
2. PCR: the cultured E.coli cells. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Genomic DNA extraction: extracting genomic DNA of E.coli.
2. Electrophoresis: gel is bad. 　　　

Person:Wenjun Wang 　　　

Experiment:
1. Agarose electrophoresis: detect the plasmid PCR product.
2. PCR: Get mamI sequence from PCR product.
3. PCR: Get mamL sequence from PCR product.
4. PCR: Get mamK sequence from PCR product to finish the Over-Lap PCR.
5. Agarose electrophoresis: detect the plasmid PCR product.
6. Product purification: purify the PCR product. 　　　

Person:Yu Wang 　　　

Experiment:
1. Pick bacteria and PCR examining, the aim fragment is 1409bp.
2. Sequencing. 　　　

July 25th

Person:Wenjun Wang 　　　

Experiment:
1. PCR: Try to get mamAB gene cluster sequence from AMB-1 bacterial strain.
2. Get the genome of AMB-1 with TaKaRa MINI Best bacteria genome DNA Extraction Ver2.0.
3. PCR: Try to get mamAB gene cluster sequence from AMB-1 genome.
4. Agarose electrophoresis: detect the plasmid PCR product. 　　　

Person:Xue Sun 　　　

Experiment:
1. d1 to d6 as primer PCR, get part K1059005.
2. Purification.
3. Recombine PCR product to pMD-19 T vector.
4. Transformation. 　　　

Person:Kaili Qin 　　　

Experiment:
1.Equip the medium：Equip the medium of AMB-1. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. SDS page gel electrophoresis: PCR production and the genomic DNA. 　　　

Person:Xiaodong Zhong 　　　

Experiment:
1. Transformation of E.coli (top10):
Part: mamK+T vector
Part: mamI+T vector
Part: mamL+T vector
Part: mamQ+T vector
Part: mamB+T vector (failure) 　　　

July 26th

Person:Yu Wang 　　　

Experiment:
1. Blue-white selection.
2. PCR examining, the aim fragment is 388bp.
3. Sequencing. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Bacterination: bacterinate PSB1C3-RFP into tubes of 10ml. 　　　

July 27th

Person:Qiu Wang 　　　

Experiment:
1. Measuring Growth curve, E.coli with RFP, pSB1C3 as plasmid backbone. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Extraction of genomic DNA from E.coli.
2. SDS page gel electrophoresis: the genomic DNA. 　　　

July 28th

Person:Qiu Wang 　　　

Experiment:
1. Measure Growth curve. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Bacterination: bacterinate PSB1C3-RFP, E.coil with single-1 protection sequence into tubes of 10ml. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Culture the E.coil cells: PSB1C3. 　　　

July 29th

Person:Yu Wang 　　　

Experiment:
1. Bacterial culturing. 　　　

Person:Wenjun Wang 　　　

Experiment:
1. PCR: Get mamB sequence from AMB-1 bacterial strain.
2. PCR: Get mamJ sequence from AMB-1 bacterial strain.
3. PCR: Get mamE sequence from AMB-1 bacterial strain.
4. PCR: Get mamE sequence from AMB-1 bacterial strain.
5. Agarose electrophoresis: detect the DNA digestion product. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Plasmid miniprep: plasmid Extraction of PSB1C3-RFP, E.coil with single-1 protection sequence for 2 tubes each.
2. Enzyme digestion: enzyme digestion of PSB1C3 and E.coil with single-1 protection sequence for 25ul system of 2 tubes each, use EcoR1, Pst1.
3. Electrophoresis: gel is good. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Miniprep: Psb1c3.
2. Digestion: digest the plasmid with EcoRI and Pst-HF.
3. SDS page gel electrophoresis.
4. Gel extraction of DNA. 　　　　　

July 30th

Person:Qiu Wang 　　　

Experiment:
1. Lstwenmin1 to Lstwenmin4 as primer PCR, get part K1059006.
2. Purification.
3. Recombine PCR product to pMD-19 T vector.
4. Transformation. 　　　　　

Person:Kaili Qin 　　　

Experiment:
1. Transform: transform the ligastion (single-1 protection sequence-PSB1C3) product into Top 10.
2. Bacterial coating. 　　　

July 31st

Person:Xue Sun 　　　

Experiment:
1. Blue-white selection.
2. PCR and electrophoresis examining, the aim fragment is 388bp and 210.
3. Sequencing.
4. The PCR result a fragment at 120bp, so we failed the part k1059006. We designed another two primers and be ready for more PCR. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Pick bacteria: pick bacteria for PCR.
2. PCR: use PCR to detect the bacteria we want (single-1 protection sequence-PSB1C3)), 25ul system. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Glycerol stocks: Psb1c3, 2 tubes.
2. Miniprep: Psb1c3.
3. Digestion: digest the plasmid with EcoRI and Pst-HF.
4. Gel extraction of DNA. 　　　

August 1st

Person:Yu Wang 　　　

Experiment:
1. Bacterination.
2. E.coil with part K1059005 PMD-19 T vector as backbone. 　　　

Person:Xiaodong Zhong 　　　

Experiment:
1. PCR genome for mamB, mamJ, mamE, mamC gene. 　　　

Person:Wenjun Wang 　　　

Experiment:
1. PCR: Get mamC sequence from AMB-1 bacterial strain.
2. Agarose electrophoresis: detect the mamC PCR product.
3. Product purification: purify the PCR product.
4. PCR: Get the mamC sequence which is more nicety from the product of product purification. 　　　

August 2nd

Person:Xue Sun 　　　

Experiment:
1. Plasmid minprep.
2. Enzyme digestion.
3. Recombination: Get part K1059005-B0035-GFPlva-K1059004-B0015, pSB1C3 as backbone. 　　　

Person:Xiaodong Zhong 　　　

Experiment:
1. Transformation of E.coil (top10): mamB+T vector. 　　　

Person:Wenjun Wang 　　　

Experiment:
1. PCR：Get mamB, mamE sequence from AMB-1 bacterial strain.
2. Get the genome of AMB-1.
3. Transform: transform the ligastion product into Top 10. 　　　

August 3rd

Person:Qiu Wang 　　　

Experiment:
1. Transformation and Bacterination. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Miniprep: Psb1c3, 6 tubes.
2. Digestion: digest the plasmid with EcoRI and Pst-HF, XbaI and Pst-HF.
3. SDS page gel electrophoresis.
4. Gel extraction of DNA. 　　　

Person:Xiaodong Zhong 　　　

Experiment:
1. E. coli Cell Culture: Top10 (mamB+T vector) Resistance: Ampicillin Medium: LB.
2. mamK PCR purification: Get mamK gene.
3. PCR genome for mamB: 16x50ul PCR system. Get mamB gene. 　　　

Person:Wenjun Wang 　　　

Experiment:
1. Agarose electrophoresis: detect the PCR product.
2. Get the genome of AMB-1.
3. Agarose electrophoresis: detect the genome product.
4. PCR: to detect the product of transform.
5. PCR: Get mamC sequence from AMB-1 bacterial strain.
6. PCR: Get 16srDNA sequence from AMB-1 bacterial strain to detect the strain.
7. Agarose electrophoresis: detect the genome product. 　　　

August 4th

Person:Kaili Qin 　　　

Experiment:
1. Ligation: J23101 order vector with chloramphenicol resistance; double2 order 　　 RBS-GFPLVA- single2-terminator and vector with chloramphenicol resistance.
2. Transform: transform the ligastion product into Top 10.
3. Bacterial coating 　　　

Person:Yu Wang 　　　

Experiment:
1. PCR: Get BBa_K1059006 sequence from PCR product.
2. Agarose electrophoresis: detect the PCR product.
3. Product purification: purify the PCR product.
4. Ligation: recombine PCR product to pMD-19 T vector.
5. Sequencing
6. Transformation: transform the ligastion product into Top 10. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. SDS page gel electrophoresis. 　　　

August 5th

Person:Qiu Wang 　　　

Experiment:
1. Blue -white selection.
2. PCR and electrophoresis examining.
3. PCR and electrophoresis examining, get the aim fragment.
4. Sequencing. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Culture the E.coli cells: pSB1C3. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Pick bacteria: pick bacteria for PCR.
2. PCR: use pcr to detect the bacteria we want.(double2-RBS-GFPLVA-single2-terminator-PSB1C3), 25ul system.
3. Electrophoresis.
4. Send sequencing.
5. Conservation: conservate E.coli with.double2-RBS-GFPLVA-single2-terminator-PSB1C3 plasmid. 　　　

August 6th

Person:Xue Sun 　　　

Experiment:
1. Plasmid minprep.
　　　

Person:Qianyun Lu 　　　

Experiment:
1. Miniprep: pSB1C3.
2. Digestion: digest the plasmid with XbaI and Pst-HF.
3. SDS page gel electrophoresis.
4. Gel extraction of DNA.
5. PCR production purification: J23106+RBS, mamQ, mamL, B0015.
6. SDS page gel electrophoresis: J23106+RBS, mamQ, mamL, B0015.

100bp, J23106+RBS, mamQ, mamL, B0015
7. PCR amplification: J23106+RBS, B0015.
8. SDS page gel electrophoresis: J23106+RBS, B0015. 　　　

Person:Wenjun Wang 　　　

Experiment:
1. PCR: Get mamJ sequence from PCR product and add prefix and suffix onto the sequence.
2. PCR: Get mamQ sequence from PCR product and add prefix and suffix onto the sequence.
3. Agarose electrophoresis: detect the PCR product.
4. Product purification: purify the PCR product.
5. Ligation: BBa_J23101-protection order 1-RBS-GFPLVA-terminator and vector with chloramphenicol resistance.
　　　

Person:Kaili Qin 　　　

Experiment:
1. Plasmid miniprep: plasmid Extraction of pSB1C3 -RFP for 5 tubes.
2. Enzyme digestion: enzyme digestion of pSB1C3 for 50ul system of 3 tubes, use Xba1, Pst1.
3. Electrophoresis: gel is good.
　　　

August 7th

Person:Xue Sun 　　　

Experiment:
1. Bacterination. 　　　

Person:Wenjun Wang 　　　

Experiment:
1. Transform: transform the ligastion product into Top 10.
2. PCR: Get mamL sequence from PCR product and add prefix and suffix onto the sequence.
3. PCR: Get mamQ sequence from PCR product and add prefix and suffix onto the sequence.
4. Agarose electrophoresis: detect the PCR product.
5. PCR: Get J23106+B0032 sequence from PCR product and add prefix and suffix onto the sequence.
6. PCR: Get B0015 sequence from PCR product and add prefix and suffix onto the sequence.
7. Agarose electrophoresis: detect the PCR product.
8. Product purification: purify the PCR product.
　　　

Person:Kaili Qin 　　　

Experiment:
1. Gel recovery: gel recovery of pSB1C3.
2. Electrophoresis: gel is good for 30ng/ul each.
3. PCR: use PCR to B0015 cloning, 50ul system.
4. Electrophoresis: gel is good.
5. PCR Purification: PCR Purification of B0015.
6. Electrophoresis: gel is good for 30ng/ul each.
7. Enzyme digestion: enzyme digestion of RBS-mamL for 25ul system, use Xba1, Pst1;RBS-mamL for 25ul system, use Spe1; B0015 for 25ul system, use Xba1; promoter-RBS for 25ul system, use EcoR1, Pst1.
8. Ligation: RBS-mamL order pSB1C3; promoter-RBS order pSB1C3; RBS-mamL order B0015.
9. Transform: transform the ligastion product into Top 10.
10. Bacterial coating.
11. Bacterination: bacterinate E.coli with PSB1C3-RFP into tubes of 10ml. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. PCR amplification: mamL, three tubes.
2. PCR production purification: mamL, three tubes; J23106+RBS, three tubes.
3. SDS page gel electrophoresis: mamL, three tubes; J23106+RBS, three tubes.

J23106+RBS, mamL
4. Digestion: digest RBS+ mamL with XbaI and Pst-HF, digest RBS+mamL with SpeI, digest B0015 with XbaI, digest J23106+RBS with EcoRI and Pst-HF.
5. SDS page gel electrophoresis.
6. Ligation: ligate RBS+mamL with Psb1c3, ligate J23106+RBS with pSB1C3,ligate RBS+ mamL with B0015. 　　　

August 8th

Person:Qiu Wang 　　　

Experiment:
1. Miniprep the pSB1C3, J23101, RGL-K1059005. 　　　

August 9th

Person:Yu Wang 　　　

Experiment:
1. Enzyme digestion: BBa_J23101 with EcoR I and SpeI, BBa_ K1059002 with XbaI and PstI, BBa_K1059005 with EcoR I and PstI, pSB1C3 with EcoRI and PstI.
2. Gel extraction pSB1C3 as vector. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Miniprep: pSB1C3, ten tubes.
2. Digestion: three tubes are digested with XbaI and Pst-HF, two tubes are digested with EcoRI and Pst-HF.
3. SDS page gel electrophoresis.
4. Gel extraction of DNA.
5. PCR amplification: mamQ, four tubes.
6. SDS page gel electrophoresis.
7. PCR production purification: mamQ, two tubes.
8. SDS page gel electrophoresis.
9. Digestion: digest J23106+RBS with EcoRI and SpeI, digest RBS+mamQ with XbaI and Pst-HF.
10. PCR amplification: RBS+mamL, four tubes. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Pick bacteria: pick bacteria for PCR.
2. PCR: use pcr to detect the bacteria we want(promoter-RBS-PSB1C3, RBS-mamL-PSB1C3), 25ul system.
3. Enzyme digestion: enzyme digestion of PSB1C3 for 50ul system, 3 tubes, use Xba1, Pst1; pSB1C3 for 25ul system of 2 tubes, use EcoR1, Pst1.
4. PCR: use pcr to cloning RBS-mamL-B0015, 50ul system.
5.Electrophoresis: gel is good.
6. Gel recovery: gel recovery of pSB1C3.
7. Electrophoresis: gel is good for 20ng/ul each.

Electrophoresis of PSB1C3 vector
　　　

August 10th

Person:Xue Sun 　　　

Experiment:
1. Recombination. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. SDS page gel electrophoresis.
2. PCR amplification: J23106+RBS, two tubes; B0015, two tubes.
3. PCR production purification: RBS+mamL, four tubes.
4. SDS page gel electrophoresis.
5. Digestion: digest RBS+mamL with SpeI.
6. Culture the E.coli cells: JB11.
7. PCR production purification: mamI.
8. SDS page gel electrophoresis. 　　　

Person:Wenjun Wang 　　　

Experiment:
1. Product purification: purify the PCR product, J1.
2. Plastid extraction: B0015, J2310, C+.
3. Digestion: use enzyme XbaI, Pst I to get part B0015, a double terminator.
4. Ligation: mamK onto pMB-19 T, prepare for sequence.
5. PCR: amplification mamJ sequence from PCR product.
6. Agarose electrophoresis: detect the PCR product.
7. Get part for agrose, mamJ.
8. PCR: Get mamI sequence from PCR product and add prefix and suffix onto the sequence.
9. Agarose electrophoresis: detect the PCR product.
10. Product purification: purify the PCR product. 　　　

Person:Kaili Qin 　　　

Experiment:
1. PCR Purification: PCR Purification of mamL1~4.
2. Conservation: promoter-RBS-PSB1C3 (P4), RBS-mamL-PSB1C3 (L4).
3. Ligation: RBS-mamJ, RBS-mamI, RBS-mamB order PMD-19 vector separately. 　　　

August 11th

Person:Yu Wang 　　　

Experiment:
1. Transformation.
2. Plate cultivation for 14h. 　　　

Person:Kaili Qin 　　　

Experiment:
1. PCR: use PCR to detect mamQ1~10, 25ul system.
2. Electrophoresis: gel is bad.
3. Transform: transform the ligastion product ( RBS-mamJ-T vector;RBS-mamI-T vector, RBS-mamB-T vector, RBS-mamK-T vector) into Top 10. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Miniprep: JB11, B0015, each two tubes.
2. Digestion: digest JB11 with EcoRI and SpeI, digest B0015 with XbaI and Pst-HF, digest RBS+mamL with XbaI and SpeI.
3. Ligation: ligate RBS+J23106 with pSB1C3, ligate RBS+mamL and BOO15 with pSB1C3. 　　　

August 12th

Person:Xue Sun 　　　

Experiment:
1. Pick bacteria and PCR examining 　　　

Person:Kaili Qin 　　　

Experiment:
1. Transform: transform the ligastion product(RBS-mamL-B0015-PSB1C3; promoter-RBS-PSB1C3) into Top 10.
2. Bacterial coating.
3. Pick bacteria: pick bacteria we want(RBS-mamL-B0015-PSB1C3, promoter-RBS-mamQ-PSB1C3). 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Pick the bacteria and culture the E.coli cells for four hours.
2. PCR: the cultured E.coli cells. 　　　

August 13th

Person:Qianyun Lu 　　　

Experiment:
1. PCR: the cultured E.coli cells.
2. SDS page gel electrophoresis.
3. Ligation: ligate the purified mamB and mamL with T vector.
4. PCR amplification: mamB, mamK, mamJ.
5. SDS page gel electrophoresis. 　　　

Person:Qiu Wang 　　　

Experiment:
1. Sequencing. 　　　

August 14th

Person:Qianyun Lu 　　　

Experiment:
1. PCR amplification: mamJ.
2. SDS page gel electrophoresis. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Pick bacteria: pick bacteria we want(RBS-mamL-B0015-PSB1C3, promoter-RBS-mamQ-PSB1C3). 　　　

Person:Yu Wang 　　　

Experiment:
1. Bacterination.
2. Make competent cells. 　　　

August 15th

Person:Wenjun Wang 　　　

Experiment:
1. PCR: get mamJ sequence from AMB-1 bacterial strain.
2. Agarose electrophoresis: detect the plasmid PCR product.
3. Product purification: purify the PCR product.
4. PCR: get mamC sequence from AMB-1 bacterial strain. 　　　

Person:Qiu Wang 　　　

Experiment:
1. Miniprep.
2. Make gel. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. SDS page gel electrophoresis.
2. PCR amplification: mamL.
3. PCR production purification: RBS+mamL.
4. SDS page gel electrophoresis. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Ligastion: mamJ, mamK order PMD19-T vector. 　　　

August 16th

Person:Xue Sun 　　　

Experiment:
1. Enzyme digestion. 　　　

Person:Kaili Qin 　　　

Experiment:
1. PCR: use PCR to detect mamK-pMD19-T vector1~15, 25ul system. 　　　

Person:Qianyun Lu

Experiment:
1. PCR amplification: mamL.
2. SDS page gel electrophoresis. 　　　

August 17th

Person:Wenjun Wang

Experiment:
1. PCR: amplification mamC sequence from PCR product.
2. Agarose electrophoresis: detect the plasmid PCR product.
3. Product purification: purify the PCR product.
4. PCR: amplification mamI sequence from PCR product.
5. PCR: amplification mamJ sequence from PCR product.
6. Agarose electrophoresis: detect the plasmid PCR product.
7. Product purification: purify the PCR product.
8. PCR: amplification mamB sequence from PCR product.
9. Agarose electrophoresis: detect the plasmid PCR product.
10. Product purification: purify the PCR product. 　　　

Person:Qianyun Lu

Experiment:
1. SDS page gel electrophoresis.
2. PCR amplification: mamQ, two tubes.
3. PCR production purification: RBS+mamL.
4. SDS page gel electrophoresis.
5. PCR amplification: mamK, two tubes; mamB, two tubes.
6. SDS page gel electrophoresis. 　　　

Person:Kaili Qin

Experiment:
1. Electrophoresis: mamK-PMD19-T vector1~15, gel is bad.
2. Pick bacteria: pick bacteria we want (mamK-PMD19-T vector1~10).
3. PCR: use PCR to detect mamK-pMD19-T vector1~10, 25ul system. 　　　

Person:Xue Sun

Experiment:
1. Recombination. 　　　

August 18th

Person:Yu Wang 　　　

Experiment:
1. Transformation.
2. Plate cultivation for 14h. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Gel recovery: gel recovery of mamE.
2. Electrophoresis: gel is good for 8ng/ul each.
3. Bacterination: bacterination of E.coli with PSB1C3 plasmid for 3 tubes, 10ml each.
4. Bacterial coating: JQ11, QP11, KP1, BP1, BP2, LP21.
5. Gel recovery: gel recovery of PSB1C3 vector.
6. Electrophoresis: gel is good for 20ng/ul each.

Electrophoresis of PSB1C3 vector
　　　

Person:Qianyun Lu 　　　

Experiment:
1. PCR amplification: mamI, two tubes; mamQ, two tubes.
2. SDS page gel electrophoresis.
3. PCR production purification: RBS+mamQ, RBS+mamI.
4. SDS page gel electrophoresis.
5. PCR amplification: mamI, two tubes; mamJ, two tubes.
6. SDS page gel electrophoresis.
7. Transformation: JQ11, QP11, BP1, BP2, LP21, KP1.
8. SDS page gel electrophoresis.
9. PCR amplification: mamI, two tubes; mamB, two tubes.
10. SDS page gel electrophoresis: mamK; mamB.

lane6 mamK1; lane7 mamK2; lane8 mamB
　　　

Person:Wenjun Wang 　　　

Experiment:
1. Digestion: use enzyme EcoR I, Pst I to Cut PSB1C3 plasmid vector.
2. Digestion: use enzyme EcoR I, Pst I to Cut promoter J23106 and RBS B0032.
3. PCR: amplification mamE, mamJ sequence from PCR product.
4. Agarose electrophoresis: detect the plasmid PCR product.
5. Ligastion: promoter, RBS and mamQ.
6. Ligastion: mamQ and PSB1C3.
7. Ligastion: mamB and PSB1C3.
8. Ligastion: mamL and PSB1C3.
9. Ligastion: mamK and PSB1C3.
10. Transform: transform the ligastion product into Top 10. 　　　

August 19th

Person:Xue Sun 　　　

Experiment:
1. Pick bacteria and PCR examining. 　　　

Person:Kaili Qin 　　　

Experiment:
1. PCR: use PCR to detect mamK-PSB1C3 vector1~10,25ul system.
2. Electrophoresis: gel is bad. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Ligation: ligate the purified mamE with T vector.
2. PCR production purification: RBS+mamB, two tubes.
3. Pick the bacteria and culture the E.coli cells for four hours. 　　　

Person:Wenjun Wang 　　　

Experiment:
1. PCR: amplification mamE sequence from PCR product.
2. Agarose electrophoresis: detect the plasmid PCR product.
3. PCR: amplification mamI sequence from PCR product.
4. Product purification: purify the PCR product.
5. Agarose electrophoresis: detect the PCR product purification.
　　　

August 20th

Person:Qianyun Lu 　　　

Experiment:
1. Pick the bacteria and culture the E.coli cells for four hours.
2. PCR production purification: RBS+mamI, two tubes; RBS+mamB, one tube.
3. SDS page gel electrophoresis.
4. PCR amplification: mamI, four tubes.
5. SDS page gel electrophoresis.
6. Pick the bacteria and culture the E.coli cells for four hours. 　　　

Person:Qiu Wang 　　　

Experiment:
1. Sequencing J23101. 　　　

Person:Kaili Qin 　　　

Experiment:
1. PCR: use PCR to detect promoter-RBS-mamQ-PSB1C3 vector1~10, 25ul system; mamK - PSB1C31~28, 25ul system; mamE-PMD19-T vector1~8, 25ul system.
2. Electrophoresis: gel is bad. 　　　

August 21st

Person:Qianyun Lu 　　　

Experiment:
1. SDS page gel electrophoresis.
2. Miniprep: pSB1C3.
3. Digestion: three tubes are digested with XbaI and Pst-HF, two tubes are digested with EcoRI and Pst-HF.
4. SDS page gel electrophoresis. 　　　

Person:Kaili Qin 　　　

Experiment:
1. PCR: use PCR to clone mamL standard, mamB standard, mamQ standard, mamK standard.
2. Electrophoresis: mamL, K, B gel is good; mamQ, gel is bad.

Electrophoresis of production of mamK,B cloning
　　　

Person:Yu Wang 　　　

Experiment:
1. Bacterial activation and culturing, including control group and all experimental groups.
2. Preparing for measuring the fluorescence. 　　　

August 22th

Person:Wenjun Wang 　　　

Experiment:
1. PCR: Get mamQ sequence from PCR product and add prefix and suffix onto the squence.
2. Agarose electrophoresis: detect the plasmid PCR product.
3. Product purification: purify the PCR product. 　　　

Person:Qiu Wang 　　　

Experiment:
1. Bacterialculturing. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. PCR amplification: mamB, four tubes; mamK, four tubes.
2. SDS page gel electrophoresis.
3. Gel extraction of DNA. 　　　

Person:Kaili Qin 　　　

Experiment:
1. PCR Purification: PCR Purification of mamL, K, B.
2. Enzyme digestion: enzyme digestion of mamL, K, B for 25ul system separately, use Xba1, Pst1. 　　　

August 23rd

Person:Xue Sun 　　　

Experiment:
1. Measuring the fluorescence. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Electrophoresis: production of mamL, K, B enzyme digestion, gel is good.
2. Ligastion: mamL, K, B order pSB1C3 vector with chloramphenicol resistance.
3. Transform: transform the ligastion product into Top 10.
4. Bacterial coating. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Ligation: ligate RBS+ mamB with pSB1C3, ligate RBS+mamL and BOO15 with pSB1C3, ligate RBS+ mamK with pSB1C3.
2. PCR amplification: mamB, four tubes.
3. Transformation: L-P, B-P, K-P.
4. SDS page gel electrophoresis.
5. Ligation: ligate the purified mamC and mamE with T vector.
6. Transformation: C-T, E-T. 　　　

Person:Wenjun Wang 　　　

Experiment:
1. PCR: Get mamI sequence from PCR product and add prefix and suffix onto the sequence.
2. PCR: Get mamL sequence from PCR product and add prefix and suffix onto the sequence.
3. PCR: Get mamB sequence from PCR product and add prefix and suffix onto the sequence.
4. Agarose electrophoresis: detect the plasmid PCR product.
5. Product purification: purify the PCR product. 　　　

August 24th

Person:Wenjun Wang 　　　

Experiment:
1. PCR: amplification mamB+mamL, mamQ+mamI sequence from DNA ligastion product.
2. Agarose electrophoresis: detect the plasmid PCR product.
3. PCR: Get mamI sequence from AMB-1 bacterial strain.
4. PCR: Get mamI sequence from PCR product and add prefix and suffix onto the sequence.
5. Product purification: purify the PCR product.
6. Agarose electrophoresis: detect the plasmid PCR product. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Pick the bacteria and culture the E.coli cells for four hours.
2. PCR: C-T, E-T, L-P, B-P, K-P.
3. SDS page gel electrophoresis: PCR production.
　　　

Person:Kaili Qin 　　　

Experiment:
1. Pick bacteria: pick bacteria we want (mamL, K, B-PSB1C3).
2. PCR: use PCR to detect mamL, K, B-PSB1C3 vector1~10, 25ul.
3. Electrophoresis: gel is bad. 　　　

Person:Xue Sun 　　　

Experiment:
1. Result analysis. 　　　

August 25th

Person:Yu Wang 　　　

Experiment:
1. Bacterial culturing, including control group and all experimental group. 　　　

Person:Kaili Qin 　　　

Experiment:
　　1. Ligastion: mamC, mamE order pMD19-T vector.
2. Transform: transform the ligastion product into Top 10.
3. Bacterial coating. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. PCR: L-P.
2. SDS page gel electrophoresis: mamC; mamE.

lane5: mamC, lane6: mamE
　　　

Person:Wenjun Wang 　　　

Experiment:
1. PCR: Get mamJ sequence from AMB-1 bacterial strain.
2. Digestion: use enzyme XbaI I, Spel I to cut mamI, mamL, mamB, mamQ.
3. Plasmid Extraction: extract the pSB1C3 plasmid which containing B0015 from E.coli.
4. Agarose electrophoresis: detect the plasmid extraction product and PCR product.
5. Ligastion: mamB and mamL; mamQ and mamI.
6. Agarose electrophoresis: detect the DNA digestion product.
7. Bacterial inoculation: inoculate the E.coli which contain pSB1C3 vector plasmid, C+.
8. PCR: amplification mamB+mamL, mamQ+mamI sequence from DNA ligastion product.
9. Agarose electrophoresis: detect the plasmid PCR product. 　　　

August 26th

Person:Xue Sun 　　　

Experiment:
1. Plasmid minprep.
2. Enzyme digestion and electrophoresis examining.
3. Recombination. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Pick bacteria: pick bacteria we want (mamC, E-pMD19T- vector).
2. PCR: use PCR to detect mamC, E-pMD19T- vector1~10, 25ul.
3. Electrophoresis: gel is bad.
4. Pick bacteria: pick bacteria we want(mamC,E-PMD19T- vector ). 　　　

Person:Qianyun Lu 　　　

Person:Yu Wang 　　　

Experiment:
1. Plasmid minprep.
2. Enzyme digestion and electrophoresis examining.
3. Recombination. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Pick bacteria: pick bacteria we want (mamL, pSB1C3 vector ).
2.Transform: transform promoter-RBS-mamB, promoter-RBS-mamQ, mamK-PSB1C3 into Top10.
3. PCR: use PCR to detect mamK-PSB1C3, 25ul system. 　　　

September 2nd

Person:Xue Sun 　　　

Experiment:
1. Transformation. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Electrophoresis: mamL1~20, 3, 12, 14. gel is good.

Electrophoresis of mamL
2. Pick bacteria: pick bacteria we want (promoter-RBS-mamB, promoter-RBS-mamQ).
3. PCR: use PCR to detect promoter-RBS-mamB, promoter-RBS-mamQ. 　　　

September 3rd

Person:Qiu Wang 　　　

Experiment:
1. Grow bacteria in the culture plates. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Bacterial coating: mamK-PSB1C3. 　　　

September 4th

Person:Kaili Qin 　　　

Experiment:
1. Pick bacteria: pick bacteria we want (mamK-PSB1C3).
2. PCR: use PCR to detect (mamK-PSB1C3).
3. Electrophoresis: mamK-PSB1C3 number5, gel is good, others are bad. 　　　

Person:Yu Wang 　　　

Experiment:
1. Pick up the single colony, PCR, run gel and sequencing.
2. Miniprep of B0015 and four experiment groups.
3. Cultured cells of PSB1C3 for making the backbone. 　　　

September 5th

Person:Qiu Wang 　　　

Experiment:
1. Miniprep the PSB1C3.
2. Enzyme digestion: B0015 with EcoR I and SpeI; the experiment group 1 use XbaI and PstI; the other experiments group use EcoR I and XbaI; the PSB1C3 with EcoRI and PstI.
3. Gel extraction PSB1C3 and the experiment group 1.
4. Recombination. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Pick bacteria: pick bacteria we want (mamK-PSB1C3).
2. PCR: use PCR to detect (mamK-PSB1C3).
3. Electrophoresis: gel is bad. 　　　

September 6th

Person:Xue Sun 　　　

Experiment:
1. Transformation. 　　　

September 9th

Person:Yu Wang 　　　

Experiment:
1. Pick up single colony, PCR, and sequencing. 　　　

Person:Kaili Qin 　　　

Experiment:
1. PCR: use PCR to clone mamC.
2. Electrophoresis: gel is bad.
3. Enzyme digestion: enzyme digestion of mamI, mamQ; 25ul system, use Xba1, Pst1.
4. Electrophoresis: gel is good. 　　　

September 10th

Person:Xue Sun 　　　

Experiment:
1. Culture the experiment group 1. 　　　

Person:Kaili Qin 　　　

Experiment:
1. Pick bacteria: pick bacteria we want (mamQ1~40, mamI1~10).
2. PCR: use PCR to detect mamQ, mamI.
3. Electrophoresis: gel is good.

Electrophoresis of mamI
　　　

September 11st

Person:Qiu Wang 　　　

Experiment:
1. Miniprep and enzyme digestion with EcoR I and XbaI.
2. Gel extraction. 　　　

September 12th

Person:Kaili Qin 　　　

Experiment:
1. Bacterination: Bacterinate E.coli with pSB2K3-RFP into tubes of 10ml. 　　　

Person:Xiaodong Zhong 　　　

Experiment:
1. TEM (Electron microscopy) of Magnetospirillum Magneticum AMB-1.

　　　

September 13th

Person:Kaili Qin 　　　

Experiment:
1. Plasmid miniprep: plasmid extraction of pSB2K3 for 5 tubes.
2. Enzyme digestion: enzyme digestion of pSB2K3 for 50ul system, use EcoR1, Pst1.
3. Gel recovery: gel recovery of the product of enzyme digestion.
4. Electrophoresis: gel is good. 　　　

September 14th

Person:Xue Sun 　　　

Experiment:
1. Inoculate the cell to culture plates, containing the control group, experiment 3 and experiment 4. 　　　

September 15th

Person:Yu Wang 　　　

Experiment:
1. Fluorescence detection. 　　　

Person:Qianyun Lu 　　　

Experiment:
1. Microscopy of Magnetospirillum magneticum AMB-1.

　　　

Person:Xiaodong Zhong 　　　

Experiment:
1. Magnetic analysis: detect the magnetism of Magnetospirillum magneticum AMB-1.

September 20th

Person:Kaili Qin 　　　

Experiment:
1. Plasmid miniprep: plasmid Extraction of double expression plasmid for 9 tubes.
2. Electrophoresis: gel is good.

Electrophoresis of double expression plasmid
　　　

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Team:OUC-China/Lab note

From 2013.igem.org

Revision as of 08:29, 26 September 2013

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