Team:NEU CHINA/Note Book
Notebook
May
| Date | Content |
|---|---|
| 2021/5/25 |
1. Sterilize a bottle of PBS 2. Compound LBA fluid medium 300ml solid medium 200ml LB fluid medium 300ml solid medium 200ml 3.Bacteria recovery Bl21,DH5α,control(5ml LB) |
| 2021/5/26 |
1.Prepare experimental equipment(PCR tube, 1.5ml ep tube, 250mlConical flask, 50mlCentrifuge tube, Petri dishes) 2.Preparation of competent cells |
| 2021/5/27 | 1.Transformation mp9 (dilute from 57ng/ul to 10ng/ul, total volume50ul, 9ul mp9+ 41ul sterile water) |
| 2021/5/28 | 1.Prepare experimental material (LBK, LBC) |
| 2021/5/29 |
1.Pick the bacteria, PCR Amplification, 2.CompoundLBA solid medium300ml, fluid medium300ml 3.mp4 Transformed into LBK solid medium |
| 2021/5/30 |
1.mp9 Double Digestion, Measure the concentration 2.Prepare experimental equipment (250mlConical flask, Centrifuge tube) 3.Prepare experimental material(DH5α, control) 4.Pick the bacteriamp3, mp9(Bl21), pacyc |
| 2021/5/31 |
1.Preparation of competent cells 2.Plasmid Extractionmp3, pacyc |
June
| Date | Content |
|---|---|
| 2021/6/1 |
1.Double Digestion mp3 2.EGFP for PCR, cycle pure |
| 2021/6/2 |
1.Prepare experimental equipment(PCR tube, 1.5ml ep tube, 250mlConical flask, 50mlCentrifuge tube, Petri dishes) 2.Preparation of competent cells |
| 2021/6/3 |
1.Plasmid ExtractionPUC57 2.PCR Amplification luxI, egfp 3.Cycle pure 4.Double Digestion, Gel Extraction, T4 connection |
| 2021/6/4 |
1.Cycle pure, Measure the concentration, Double Digestion 2.Gel Extraction, Measure the concentration 3.Connection and transformation |
| 2021/6/5 |
1.Double Digestion, Gel Extraction, Measure the concentration 2.Connection and transformation |
| 2021/6/6 |
1.Pick the bacteria, Amplification 2.Plasmid Extraction |
| 2021/6/7 |
1.Pick the bacteria, Amplification 2.Plasmid Extractionmp2-1 |
| 2021/6/8 |
1.Measure plasmid concentration. Sent to the company for sequencing 2.The plux was amplified by PCR 3.Gel Extraction, Double Digestion, Gel Extraction 4.Connection and transformation |
| 2021/6/9 |
1.Measure the concentration, connection hrp and mp2-1 2.Transformation 3.Plasmid Extraction |
| 2021/6/10 |
1.The plux was amplified by PCR 2.Pick the bacteriamp2-3, cycle pure |
| 2021/6/26 |
1.Hrp PCR Amplification from mp3 2.Cycle pure, Measure the concentration 3.Double Digestion hrp and mp2-1vector 4.Connection, Transformation 5.AGE test |
| 2021/6/27 |
1.PCR Amplification luxI and luxR 2.Cycle pure 3.Double Digestion luxI and luxR 4.Gel Extraction, Connection and transformation 5.Prepare experimental equipment and material 6.Shake the bacteria Amplification |
| 2021/6/28 |
1.Connect luxI and luxR, Gel Extraction 2.Plasmid Extraction mp1-1, Measure the concentration 3.Colony PCR 4.Digestion plasmid mp1-1and luxI-luxR 5.Connection and transformation |
| 2021/6/29 |
1.Double Digestion mp1-1 2.Connect mp1-1, luxI, luxR, Transformation 3.Connect mp131, luxI-R, Transformation 4.Amplification mp1-1 |
| 2021/6/30 | 1.Plasmid Extraction |
July
| Date | Content |
|---|---|
| 2021/7/1 | 1.Plasmid Extraction |
| 2021/7/2 |
1.AGE test mp1 2.Double Digestion mp2-1 3.PCR Amplification LuxI-LuxR, LuxI-Egfp 4.Cycle pure, Measure the concentration 5.mp8 Transformation DH5α |
| 2021/7/3 |
1.PCRAmplificationluxI-Egfp 2.Digestion pAcycduet-1 and luxI-Egfp; DigestionluxI-luxRandmp1-1 3.Gel Extraction 4.Connection and transformation 5.PCR Amplification luxI, Egfp 6.Cycle pure 7.Digestion and Connection 8.Pick the bacteria |
| 2021/7/4 |
1.PCR Amplification luxI-Egfp 2.Cycle pure 3.Digestion luxI-Egfp and pAcycduet-1 4.Gel Extraction, connection, Transformation 5.ColonyPCR, marking on the panel 6.Prepare experimental material (LBA and LBC) |
| 2021/7/5 |
1.ColonyPCR 2.AGE test 3.TvectorConnection and transformation |
| 2021/7/6 |
1.Prepare experimental material (LBC, LBA, LB, IPTG) 2.T vector connection, Transformation |
| 2021/7/7 |
1.T7 connection 2.Pick the bacteria from NO.12, 14, 15, 16 3.TransformationDH5α(mp8) |
| 2021/7/8 |
1.PCR Amplification luxI-Egfp, luxI-luxI-luxR 2.Gel Extraction 3.ColonyPCR 4.Amplification pacyc 5.Pick the bacteria, Shake the bacteria |
| 2021/7/9 |
1.Connection and transformation 2.Plasmid Extraction mp8, Measure the concentration |
| 2021/7/10 |
1.PCR Amplification luxI, Egfp 2.Cycle pure 3.luxI, Egfp, Digestion and connection 4.Gel Extraction 5. PCR Amplification 6.Shake the bacteriamp1, mp2 |
| 2021/7/11 |
1.Connect luxI-Egfp 2.PCR check problems 3.Plasmid Extraction 4.DoubleDigestion 5.Connection and transformation 6.ColonyPCR 7.Shake the bacteria (MERS,229E) |
| 2021/7/12 |
1.Shake the bacteria 2.Plasmid Extractionpuc57, mp1-1 3.TransformationPET-30a (+) 4.Prepare experimental material 5.Plasmid Extraction, Measure the concentration |
| 2021/7/13 |
1.Plasmid Extraction puc57 mp1-1 2.Overlapping PCR Amplification luxI, luxR 3.Overlapping PCR Amplification luxI-luxR 4.Digestionmp1-1, luxI-R 5.Pick 229E, MERS, Shake overnight 6.Compound PBS(PH6.2), compound lysozyme solution (10mg/ml) |
| 2021/7/14 |
1.CIP process, mp1-1 digestion 2.Gel Extraction 3.AGE verify 4.Connection and transformation 5.Plasmid Extraction 6.Protein extraction, compound Gel 7.Shake the bacteria |
| 2021/7/15 |
1.ColonyPCR 2.Preparation of competent cells 3.Compound Gel, Western Blot, Transfer, Block, Primary antibody incubation |
| 2021/7/16 |
1.Connection and transformation 2.Shake the bacteria 3.Secondary antibody incubation, Exposure |
| 2021/7/17 |
1.Digestion mp1-1 2.Gel Extraction, Measure the concentration 3.Connection and transformation, Shake the bacteria pETDuet-1 4.Smear plate MERS(Bl21) 5.Protein extraction, purify 6.Culture bacteria |
| 2021/7/18 |
1.Colony PCR 2.Shake the bacteriaDH5α 3.Transformation NO.1 and 6 4.Pick the bacteria NO.4, 5, 12, 15 5.Compound Gel, Western Blot, Primary antibody incubation |
| 2021/7/19 |
1.Preparation of competent cells 2.Pick the bacteria mp2-1(1,6) 3.Plasmid Extraction NO. 4,5,12,15 4.Digestion test 5.ColonyPCR 6.Digestion luxI-R; Gel Extraction; Connection (luxI-R：mp1-1=1：5); Transformation 7.Shake the bacteria MERS, 229E 8.Protein extraction, Ni-sepharose purify 9.Exposure |
| 2021/7/20 |
1.ColonyPCR, 1:5 2.Plasmid Extraction mp2-1 3.AGE test 4.Pick the bacteriamp1 5.Prepare experimental material (IPTG) 6.Big Protein extraction, Ni-sepharose purify 7.Shake the bacteria, measure OD, big shaker |
| 2021/7/21 |
1.Plasmid Extraction, Measure the concentration 2.Digestion, AGE test 3.Gel Extraction 4.PCR Amplification luxI-luxR 5.Connect luxI-luxR and mp2-1 6.Big 229E Extraction |
| 2021/7/22 |
1.Digestion 2 tube mp1-1 2.Gelrecycle 3.Connection and transformation 4.Western Blot; Primary antibody incubation 5.Small extract twice, big Extract once |
| 2021/7/23 |
1.Plasmid Extraction mp2-1 2.Secondary antibody incubation, Exposure |
| 2021/7/25 |
1.Shake 2 tubes of pETDeut-1 2.PCR Amplification 3.Gel recycle 4.Prepare experimental equipment |
| 2021/7/26 |
1.Shake 2 tubes of BS-united bacteria solution 2.Send BNU_China mp1-1bacteria solution 3.Plasmid extraction mp1-1, Measure the concentration 4.Digestionmp1-1, egfp 5.Gel Extraction 6.Connection and transformation 7.Plasmid Extraction BS_united_China, part-1, part-2 8.Pick two tubes of bacteria and shake them to resuscitate |
| 2021/7/27 |
1.ColonyPCR 2.Use vector primer PCR 3.Shake the bacteria 4.Resuscitate two tubes of glycerol bacteria, shake (one tube each of MERS, 229E) 5.Prepare experimental material (two bottles of LBK) |
| 2021/7/28 |
1.Plasmid Extraction tube NO.1, 4, 6, 15; Leave the sample; Smear plates; Send for sequencing 2.Measure the concentration; Line on the flat 3.Transformation Bl21 4.bacteria solution; Separate packaging; Centrifugal collect the sediment 5.Plasmid extraction; Measure the concentration |
| 2021/7/29 |
1.Transformation 2.Borrow A218 50% glycerinum, sterilization; Cryopreserve used for bacteria |
| 2021/7/30 |
1.Pick two tubes each of the bacteria MERS,229E; Cryopreserve one 2.Make a tube of 50KD parallel control |
| 2021/7/31 |
1.Cryopreserve 4 tubes of each glycerinum bacteria 2.Shake for 4h on big shaker; Add IPTG 3.Extract total Protein |
August
| Date | Content |
|---|---|
| 2021/8/1 |
1.AGE |
| 2021/8/2 |
1.Shake bacteria contain pmrB(229E),mp1-2 2.Plasmid Extraction 3.Shake mp1-2(Tube NO.6) 4.Ni-sepharose purify |
| 2021/8/3 |
1.Plasmid Extraction pUC57 contains pmrB (229E) 2.Double Digestion mp1-2, pmrB 3.Connection and transformation 1:10 4.Shake 4 tubes on small shaker 5.Wash Gel |
| 2021/8/4 |
1.ColonyPCR, transfer to big shaker 2.Pick the bacteria NO.26,8,11 3.Compound Gel, Prepare SDS+CBB dye, Add IPTG and S Protein |
| 2021/8/5 |
1.Plasmid Extraction 2.Send for sequencing 3.Compound 10 LBA 4.Repeat and shake; Add IPTG, S Protein (This time add total protein) |
| 2021/8/6 |
1.Line on the flat |
| 2021/8/7 |
1.Shake mp2-2 on small shaker 2.Shake a tube of MERS bacteria solution on small shaker overnight 3.AGE verify; CBB dye |
| 2021/8/8 |
1.Compound 200μm AHL 2.Shake on big shaker 3.Add AHL expression 4.PCRAmplificationhrp 5.Cycle pure 6.Shake the bacteria 7.Shake a bottle of MERS bacteria solution on big shaker 8.CompoundCBB dye reagent R250 and eluent 9.Exposure |
| 2021/8/9 |
1.Plasmid Extraction 2.Shake mp2-2 Tube NO.6 3.ExtractMERS Protein, BCA Measure the concentration, AGECBB dye 4.Pick Glycerin bacteria |
| 2021/8/10 |
1.Measure the growth curve; Measure the effect of AHL on thallus growth every hour 2.Compound LBK; Reverse the flat 3.Plasmid Extraction, Measure the concentration |
| 2021/8/11 |
1.Gel Extraction hrp NO.28, mp2-2 NO.32 2.Connection and transformation 3.Culture together, cryopreserve 10 tubes of sediment |
| 2021/8/13 |
1.Plasmid Extractionmp2-2 Tube NO.4 2.Hrp PCR Amplification 3.Cycle pure 4.Digestion, Gel Extractionmp2-2 Tube NO.9, hrp Tube NO.8 5.Connection and transformation |
| 2021/8/14 |
1.Digestion, Gel Extraction 2.Connection and transformation 3.Protein extraction, purify, Add PMSF for ultrasound, Ni-sepharose purify, Shake with hand 4.AGECBB dye |
| 2021/8/15 |
1.Hrp connect to mp2-2 2.LuxI connect to mp2-1 3.Colony recovery |
| 2021/8/16 |
1.Line on the flatmp2-2(with hrp), mp1, luxI (not growing) 2.PCR Amplification luxI, luxR 3.Connect mp1-1, luxI 4.Sterilize LBC solid medium, LB fluid medium 5.Shake 4 tubes of bacteria prepared for expression 6.Reverse the flat 7.Repeat and shake; Add IPTG, S Protein; See the result 8.Ni-sepharose purify recycle |
| 2021/8/17 |
1.ColonyPCR 2.Connect hrp again, Transformation 3.Pick the bacteria luxI Tube NO.1, 2, 3, 4 4.Shake the bacteria; Prepare for characterization |
| 2021/8/18 |
1.ColonyPCR hrp (no result) 2.characterization, transfer to big shaker from small shaker, after 4h, Add AHL, Measure the fluorescence OD 3.Digestionmp1-3 4.Connection and transformation |
| 2021/8/19 |
1.ColonyPCR 2.mp2-2, luxI-luxR; Pick the bacteria NO.6, 7, 12, 13 |
| 2021/8/20 |
1.Plasmid Extraction mp1 Bacteria NO.1, 2, 3, 4 2.PCR, AGE test 3.Line on the flat mp1-3, mp1 |
| 2021/8/21 |
1.Sterilize LBA, LBC 2.Transformation partⅠ,partⅡ |
| 2021/8/23 |
1.Shake the bacteria partⅠ,partⅡ,mp1(2) 2.Compound cmr |
| 2021/8/24 |
1.PartⅡcharacterization(Move to shake for 4h; Add IPTG; Measure the fluorescence, OD) 2.DigestionluxI, Connection and transformation |
| 2021/8/25 |
1.ColonyPCR 1~6,9~13 2.Pick the bacteria |
| 2021/8/26 |
1.Plasmid Extraction 2.Pick mp2-2 |
| 2021/8/27 |
1.Pick the bacteria, shake overnight |
| 2021/8/28 |
1.Plasmid Extraction Measure the concentration; Transform smear plates 2.Sterilize LBK; 50% glycerinum; Conical flask |
| 2021/8/29 |
1.Shake tube mp1-12, tube mp1(8)4, tube mp2-24 |
| 2021/8/30 |
1.Plasmid Extractionmp1, mp2-2 2.Transformation mp1 to Bl21 3.Pick the bacteria |
| 2021/8/31 |
1.Compound LBA, LBC solid medium, LB, LBK fluid medium 2.PCR Amplification hrp 3.Digestion mp2-2 4.Gel Extraction 5.Preparation of competent cells Bl21(mp1) 6.Transform mp2-2 to competent cells Bl21(mp1) |
September
| Date | Content |
|---|---|
| 2021/9/1 | 1.Pick the bacteria Shake the bacteria |
| 2021/9/2 |
1.Do characterization: transfer to big shaker, dilute OD to 0.2 2.Add AHL 3.Measure the fluorescence, OD |
| 2021/9/4 | 1.Shake the bacteria, used for characterization. The same as BS |
| 2021/9/5 |
1.9a.m ,culture for 12h,Add IPTG, S Protein partⅠ 2.Every hour have a look 3.Collect supernatant at 9 p.m |
| 2021/9/6 | 1.Measure the fluorescence, OD |
| 2021/9/7 | 1.Shake the bacteria; Purify AGE; CBB dye |
| 2021/9/8 |
1.Characterization (Give up because of infection) 2.Shake the bacteriamp1-2, mp3-2; Double transformation bacteria (mp1+mp2-2) control 3.See the stripe |
| 2021/9/9 |
1.Characterization; Transfer to big shaker for 4h,Add IPTG(1：100), S Protein(1：50), 4h see result 2.Shake the bacteria LB, LB+ competent cell, mp1-2 3 tubes (Without control; Only add IPTG, Add IPTG and S Protein), mp3-2 3 tubes |
| 2021/9/10 | 1.Characterization; Big shake for 5h, After adding IPTG and S Protein, shake for 5h, measure result |
| 2021/9/11 |
1.Digestion egfp Tube NO.1, 3, 4, pETDuct-1 2.Connection and transformation |
| 2021/9/12 | 1.ColonyPCR; Shake the bacteria on small shaker overnight |
| 2021/9/13 |
1.A set of characterization 2.Plasmid Extractionmp2-2 3.Digestion mp2-2, Gel Extraction 4.PCR Amplification hrp, cycle pure |
| 2021/9/14 |
1.ColonyPCR 2.Pick the bacteriamp3-2 Tube 3, Double Transformation, competent, LB 3.Transformation mp1-2, mp3-2 to Bl21, mp2-2 to Bl21(mp1) |
| 2021/9/15 | 1.Plasmid Extraction; Do a set of characterization, measure 100μ L every hour with a 96-well plate 2. Shake the bacteria at night |
| 2021/9/16 |
2.A set of characterization (transfer to big shaker for 4h; Add IPTG, S Protein, then measure 100μL every hour with a 96-well plate), Shake the bacteria at night 3.Transformation 3-2, Double shake |
| 2021/9/17 | 1.A set of characterization, Shake the bacteria at night |
| 2021/9/18 | 1.A set of characterization, Shake the bacteria at night |
| 2021/9/19 | 1.A set of characterization, Shake the bacteria at night |
| 2021/9/20 |
1.A set of characterization (No bacteria growing) 2.Shake the bacteria |
| 2021/9/21 | 1.A set of characterization, Shake the bacteria at night |
| 2021/9/22 |
1.A set of characterization, Shake the bacteria at night 2.Shake the bacteriamp2-2, T7-Egfp 3.Compound 9 LBA solid medium, 10 LBC solid medium 4.Plasmid Extraction T7-Egfp; Transformation to Bl21 |
| 2021/9/23 |
1.A set of characterization, Shake the bacteria at night 2.Plasmid Extraction |
| 2021/9/24 |
1.A set of characterization, Shake the bacteria at night 2.Smear plates |
| 2021/9/25 |
1.A set of characterization; Shake the bacteria at night 2.Plasmid ExtractionT7-Egfp 3.Pick the bacteria, small shaker at night |
| 2021/9/26 |
1.A set of characterization; Shake the bacteria at night 2.TransformationSARS plasmid to DH5α上Amplification 3.Protein extraction |
| 2021/9/27 |
1.A set of characterization, Shake the bacteria at night 2.AGE |
| 2021/9/28 |
1.A set of characterization, Shake the bacteria at night 2.Plasmid Extraction, Transformation |
| 2021/9/29 | 1.A set of characterization, Shake the bacteria at night |
| 2021/9/30 |
1.A set of characterization, Shake the bacteria at night 2.Cryopreserve bacteria solution |
October
| Date | Content |
|---|---|
| 2021/10/1 |
1.A set of characterization, Shake the bacteria at night |
| 2021/10/2 |
1.A set of characterization, Shake the bacteria at night |
| 2021/10/3 |
1.No bacteria growing 2.Pick the bacteria |
| 2021/10/4 |
1.A set of characterization, Shake the bacteria at night 2.Protein extraction |
| 2021/10/5 |
1.AGE 2.A set of characterization, Shake the bacteria at night |
| 2021/10/6 |
1.A set of characterization for mp9 , Shake the bacteria at night. 2.Transform. |
| 2021/10/7 |
1.A set of characterization for mp9 and transformation bacteria (mp1+mp2-2), control, Shake the bacteria at night. |
| 2021/10/7 |
1.A set of characterization for mp9 and transformation bacteria (mp1+mp2-2), control, Shake the bacteria at night. |
| 2021/10/8 |
1.A set of characterization for mp9 and transformation bacteria (mp1+mp2-2), control, Shake the bacteria at night. |
| 2021/10/9 |
1.A set of characterization for mp9 and transformation bacteria (mp1+mp2-2), control, Shake the bacteria at night. |
| 2021/10/10 |
1.A set of characterization for mp9 and transformation bacteria (mp1+mp2-2), control, Shake the bacteria at night. |
| 2021/10/11 |
1.A set of characterization for mp9 and transformation bacteria (mp1+mp2-2), control, Shake the bacteria at night. 2. Transform |
| 2021/10/12 |
1.A set of characterization for mp9 and transformation bacteria (mp1+mp2-2), control, Shake the bacteria at night. |
| 2021/10/13 |
1.A set of characterization for mp9 and transformation bacteria (mp1+mp2-2), control, Shake the bacteria at night. |
| 2021/10/14 |
1.A set of characterization for mp9 and transformation bacteria (mp1+mp2-2), control, Shake the bacteria at night. |
| 2021/10/15 |
1.A set of characterization for mp9 and transformation bacteria (mp1+mp2-2), control, Shake the bacteria at night. |