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2019.7.20 Sat

Read the literature

2019.7.21 Sun

Read the literature on arsenic treatment in Bangladesh

2019.7.22 Mon

Upon first entry into the laboratory, arsenic adsorption plasmids had been synthesized by two plasmids, pEP-SpPCS and pET-orpF-fmt, transforming E. coli BL21(DE3) and culture overnight

2019.7.23 Tue

Select a single colony on the plate, convert the colonies of pEP-SpPCS and pET-orpF-fmt, inoculate LB broth containing ampicillin resistance, and culture overnight.

According to the designed primer, J33201fragment was amplified, electrophoresis identification, gel cutting and purification, enzyme cutting E/S, gel cutting and purification

2019.7.24 Wed

The bacterial liquid cultured overnight was re-inoculated into fresh LB broth containing ambenzyl resistance at a ratio of 1:100 for expanded culture and detection of OD600. When OD value was 0.6-0.8, IPTG(1mM) was added for induction expression and low temperature was induced overnight at 28℃.

2019.7.25 Thu

Protein samples were prepared with 5* SDS-PAGE loading buffer

SDS-PAGE gel was prepared and SDS-PAGE electrophoresis was carried out on the prepared samples

After electrophoresis, Coomassie brilliant blue was dyed for 30min, then decolorized and washed overnight

2019.7.26 Fri

SDS-PAGE electrophoresis results were observed and photographed

Prepare HUST communication materials

SDS-PAGE adhesive was prepared and WB experiment was carried out. Upper sample, electrophoresis, membrane transfer, incubation of primary antibody, anti-Histag 1:10, 000,4 ℃ overnight

2019.7.27 Sat.

No experiments, participate in the HUST exchange

2019.7.28 Sun

rest

2019.7.29 Mon

SDS-PAGE adhesive was prepared and WB experiment was carried out. Upper sample, electrophoresis, membrane transfer, sealing, incubation of primary antibody, anti-Histag 1:10,000,

2019.7.30 Tue

Wash, incubate the secondary antibody, wash, expose, X-ray film, analyze the results and find that the signal is too strong, WB needs to be done again.

2019.7.31 Wed

Prepare for meetup in hangzhou. Experiment suspended

2019.8.1 Thu - 8.4. Sun

In HANGZHOU MEETUP

2019.8.5 Mon

SDS-PAGE adhesive was prepared and WB experiment was carried out.Upper sample, electrophoresis, membrane transfer, sealing, incubation of primary antibody, optimization of reaction conditions, anti-histag 1:100,000

2019.8.6 Tue

Wash, incubate the secondary antibody, wash, expose and photograph, and obtain satisfactory experimental results

2019.8.7 Wed

Plasmid containing GFP,( plate1,4K), and 10ulddH20 were found from iGEM distribution kit, and the GFP fragment was amplified as the template. Electrophoresis verification showed that the electrophoresis result was not ideal

2019.8.8 Thu

According to the redesign of primers, repcr, electrophoresis, gel cutting and purification

2019.8.9 Fri

IGEM distribution kit linearized pSB1C3 plasmid, enzyme digestion, E/P, electrophoresis validation, gel purification

J33201 fragment was cut and purified, E/S was cut and purified

The fragment of RBS-GFP was cut and purified, enzyme digestion, X/P, electrophoresis verification, and cut and purified

T4L ligase connected the above fragments at 16℃ for 1h

The junction product transformed E. coli DH5 a, coated with a plate containing chloramphenicol resistance, and cultured overnight

2019.8.10 Sat

Positive clones grew on the plate, and the plate was put in a 4℃ refrigerator for experiment on Monday

2019.8.11 Sun

Rest and do no experiments

2019.8.12 Mon

The recombinant j33201-rbs-gfp was verified by colony PCR on the plate, PCR products were electrophoresis, positive clones were selected, chloramphenicin-resistant LB broth was inoculated, and overnight culture was conducted

2019.8.13 Tue

The bacteria solution was reserved with glycerin bacteria, the remaining small plasmid was identified by enzyme digestion, and three plasmids were selected and sent to the company for sequencing

2019.8.14 Wed

Sequencing results were in line with expectations and functional measurements were initiated. Prepare arsenic preparation, prepare mother liquor

2019.8.15. Thu

Growth curve experiment

Induction curve experiment

2018.8.16 Fri

Analysis of the data

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+									<article id="post-466" class="post-466 post type-post status-publish format-standard has-post-thumbnail hentry category-standard">
+										<h2 class="entry-title"><a href="#">Notebook</a></h2>
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+											<p><br></p><p><span style=";font-family:Calibri;font-size:14px">2019.7.20 Sat </span></p><p><span style=";font-family:Calibri;font-size:14px">Read the literature</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.7.21 Sun</span></p><p><span style=";font-family:Calibri;font-size:14px">Read the literature on arsenic treatment in Bangladesh</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.7.22 Mon</span></p><p><span style=";font-family:Calibri;font-size:14px">Upon first entry into the laboratory, arsenic adsorption plasmids had been synthesized by two plasmids, pEP-SpPCS and pET-orpF-fmt, transforming E. coli BL21(DE3) and culture overnight</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.7.23 Tue</span></p><p><span style=";font-family:Calibri;font-size:14px">Select a single colony on the plate, convert the colonies of pEP-SpPCS and pET-orpF-fmt, inoculate LB broth containing ampicillin resistance, and culture overnight.</span></p><p><span style=";font-family:Calibri;font-size:14px">According to the designed primer, J33201fragment was amplified, electrophoresis identification, gel cutting and purification, enzyme cutting E/S, gel cutting and purification</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.7.24 Wed</span></p><p><span style=";font-family:Calibri;font-size:14px">The bacterial liquid cultured overnight was re-inoculated into fresh LB broth containing ambenzyl resistance at a ratio of 1:100 for expanded culture and detection of OD600. When OD value was 0.6-0.8, IPTG(1mM) was added for induction expression and low te</span><span style=";font-family:宋体;font-size:14px">mperature was induced overnight at 28<span style="font-family:宋体">℃</span><span style="font-family:Calibri">.</span></span></p><p><span style=";font-family:Calibri;font-size:14px">2019.7.25 Thu</span></p><p><span style=";font-family:Calibri;font-size:14px">Protein samples were prepared with 5* SDS-PAGE loading buffer</span></p><p><span style=";font-family:Calibri;font-size:14px">SDS-PAGE gel was prepared and SDS-PAGE electrophoresis was carried out on the prepared samples</span></p><p><span style=";font-family:Calibri;font-size:14px">After electrophoresis, Coomassie brilliant blue was dyed for 30min, then decolorized and washed overnight</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.7.26 Fri</span></p><p><span style=";font-family:Calibri;font-size:14px">SDS-PAGE electrophoresis results were observed and photographed</span></p><p><span style=";font-family:Calibri;font-size:14px">Prepare HUST communication materials</span></p><p><span style=";font-family:宋体;font-size:14px">SDS-PAGE adhesive was prepared and WB experiment was carried out.</span><span style=";font-family:Calibri;font-size:14px">&nbsp;</span><span style=";font-family:宋体;font-size:14px">Upper sample, electrophoresis, membrane transfer, incubation of primary antibody, anti-Histag 1:10, 000,4 <span style="font-family:宋体">℃ </span><span style="font-family:Calibri">overnight</span></span></p><p><span style=";font-family:Calibri;font-size:14px">2019.7.27 Sat.</span></p><p><span style=";font-family:Calibri;font-size:14px">No experiments, participate in the HUST exchange</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.7.28 Sun</span></p><p><span style=";font-family:Calibri;font-size:14px">rest</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.7.29 Mon</span></p><p><span style=";font-family:Calibri;font-size:14px">SDS-PAGE adhesive was prepared and WB experiment was carried out. Upper sample, electrophoresis, membrane transfer, sealing, incubation of primary antibody, anti-Histag 1:10,000,</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.7.30 Tue</span></p><p><span style=";font-family:Calibri;font-size:14px">Wash, incubate the secondary antibody, wash, expose, X-ray film, analyze the results and find that the signal is too strong, WB needs to be done again.</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.7.31 Wed</span></p><p><span style=";font-family:Calibri;font-size:14px">Prepare for meetup in hangzhou. Experiment suspended</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.8.1 Thu - 8.4. Sun</span></p><p><span style=";font-family:Calibri;font-size:14px">In HANGZHOU MEETUP</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.8.5 Mon</span></p><p><span style=";font-family:Calibri;font-size:14px">SDS-PAGE adhesive was prepared and WB experiment was carried out.Upper sample, electrophoresis, membrane transfer, sealing, incubation of primary antibody, optimization of reaction conditions, anti-histag 1:100,000</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.8.6 Tue</span></p><p><span style=";font-family:Calibri;font-size:14px">Wash, incubate the secondary antibody, wash, expose and photograph, and obtain satisfactory experimental results</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.8.7 Wed</span></p><p><span style=";font-family:Calibri;font-size:14px">Plasmid containing GFP,( plate1,4K), and 10ulddH20 were found from iGEM distribution kit, and the GFP fragment was amplified as the template. Electrophoresis verification showed that the electrophoresis result was not ideal</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.8.8 Thu</span></p><p><span style=";font-family:Calibri;font-size:14px">According to the redesign of primers, repcr, electrophoresis, gel cutting and purification</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.8.9 Fri</span></p><p><span style=";font-family:Calibri;font-size:14px">IGEM distribution kit linearized pSB1C3 plasmid, enzyme digestion, E/P, electrophoresis validation, gel purification</span></p><p><span style=";font-family:Calibri;font-size:14px">J33201 fragment was cut and purified, E/S was cut and purified</span></p><p><span style=";font-family:Calibri;font-size:14px">The fragment of RBS-GFP was cut and purified, enzyme digestion, X/P, electrophoresis verification, and cut and purified</span></p><p><span style=";font-family:宋体;font-size:14px">T4L ligase connected the above fragments at 16<span style="font-family:宋体">℃ </span><span style="font-family:Calibri">for 1h</span></span></p><p><span style=";font-family:Calibri;font-size:14px">The junction product transformed E. coli DH5 a, coated with a plate containing chloramphenicol resistance, and cultured overnight</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.8.10 Sat</span></p><p><span style=";font-family:宋体;font-size:14px">Positive clones grew on the plate, and the plate was put in a 4<span style="font-family:宋体">℃ </span><span style="font-family:Calibri">refrigerator for experiment on Monday</span></span></p><p><span style=";font-family:Calibri;font-size:14px">2019.8.11 Sun</span></p><p><span style=";font-family:Calibri;font-size:14px">Rest and do no experiments</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.8.12 Mon</span></p><p><span style=";font-family:Calibri;font-size:14px">The recombinant j33201-rbs-gfp was verified by colony PCR on the plate, PCR products were electrophoresis, positive clones were selected, chloramphenicin-resistant LB broth was inoculated, and overnight culture was conducted</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.8.13 Tue</span></p><p><span style=";font-family:Calibri;font-size:14px">The bacteria solution was reserved with glycerin bacteria, the remaining small plasmid was identified by enzyme digestion, and three plasmids were selected and sent to the company for sequencing</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.8.14 Wed</span></p><p><span style=";font-family:Calibri;font-size:14px">Sequencing results were in line with expectations and functional measurements were initiated. Prepare arsenic preparation, prepare mother liquor</span></p><p><span style=";font-family:Calibri;font-size:14px">2019.8.15. Thu</span></p><p><span style=";font-family:Calibri;font-size:14px">Growth curve experiment</span></p><p><span style=";font-family:Calibri;font-size:14px">Induction curve experiment</span></p><p><span style=";font-family:Calibri;font-size:14px">2018.8.16 Fri</span></p><p><span style=";font-family:Calibri;font-size:14px">Analysis of the data</span></p><p><br></p>
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Difference between revisions of "Team:Worldshaper-Wuhan/notebook"

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