Team:FAFU-CHINA/Notebook
2017 Jun
2017 Jul
2017 Agu
2017 Sep
2017 Oct
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2017.05.28-2017.06.03
We collected a lot of soil samples from all over the country, hoping to screen out the bacteria with the ability to dissolve phosphorus. Firstly, we put half of the samples into organic phosphorus and inorganic phosphorus medium for culture.
2017.06.04-2017.06.10
During the screening of bacteria, total four soil samples, respectively, 37th, 38th, Beishan area, Chinese garden soil and soil samples. Phosphorus enrichment in organic and inorganic found even filtered process 37, 38th samples did not see obvious phenomena of dissolved phosphorus, observed only yellow mucus-like plaque. Stored too long, speculation, and dry, less solubilizing bacteria, strong colony survival and stress resistance (yellow mucus-like plaque), a type of bacteria in less dissolved phosphorus under low phosphorus environment to survive. 37, 38th, soil samples failed. Found in the Beishan screening of soil samples, the only obvious solubilizing, labeled a, its propagation and flat dash found solubilizing effect, as one of candidate species for further experiments. Therefore, crossed the plate picked up three colonies, numbered A1, A2 and A3, respectively, on the inclined plane of bacteria and propagation in preparation for the molybdenum blue Colorimetry. Screening of Chinese garden soil samples found in phosphate, then pick the better of the two colonies (numbered b, c) a flat line, but the result is not ideal c, pass. B the results still can, collect a bacteria retention and propagation in preparation for molybdenum blue Colorimetry. Direct pick the bacteria found in the process of picking fungi in repeated experiments, pick the number of bacteria will not, lead to very different results.
2017.06.11-2017.06.17
Then we did the plate scribing, and picked out a single colony with phosphorus solution, measuring its 16SrDNA to identify it.
Finally, we found Bacillus megaterium.Genomic DNA was extracted from bacterial isolates using a commercial kit according to the manufacturer’s instructions. Then, DNA was stored at −20℃. Two universal oligonucleotide forward and reverse primers were synthesized based on the standard 16S rRNA gene sequence. The primers used to amplify the 16S rDNA samples were
forward primer:
-5’-CCGAATTCGTCGACAACAGAGTTTGATCCTGGCTCAG-3’.
and reverse primer:
-5’-CCCGGATCCAAGCTTACGGCTACCTTGTTACGACTT-3’.
PCR Amplification.
10x PCR Buffer 5μL
10x dNTP 0.5μL
Forward Primer 1μL
Reverse Primer 1μL
Template 1μL
Taq DNA polymerase 1μL
Sterile distill water up to 50 μL
PCR
initial denaturation at 95℃ for 3minutes which was followed with 35-cycle consisting of denaturation at ℃ for 30 seconds, annealing at 60℃ for 30 seconds, and elongation at 72℃ for 2minutes.The reaction was completed with an extension step at 72℃for 5 minutes. All samples were removed from the thermal cycler and stored at −20℃. The PCR product was determined by electrophoresis with loading 8 l of PCR product on to 1% agarose gel.
2017.06.18-2017.06.24
Considering stability and efficiency of subsequent genetic tranformantion, we bought strain from microbial and inoculum company and then cultivated the Bacillus megaterium strain from first generation to seventh generation on egg medium.
2017.06.25-2017.07.01
We continually cultivated the Bacillus megaterium strain from seventh generation to fifth generation on egg medium. However, we still didn’t find any signs of degradation of the ability to dissolve phosphorus. So we gave up the screening.
2017.07.02-2017.07.08
The growth curve of Bacillus megaterium was measured in beef extract peptone with different lead concentration. gradient
2017.07.09-2017.07.15
The growth curve of Bacillus megaterium was measured in beef extract peptone with different cadmium concentration gradient.
2017.07.16-2017.07.22
ALL of our parts were by Genscript Company, The Beijing Genomics Institute (BGI) Company and Fuzhou Sunya Biotechnology Company.2 weeks later, we got two parts.
2017.07.23-2017.07.29
2017.07.30-2017.08.05
ALL of our parts were by Genscript Company, The Beijing Genomics Institute (BGI) Company and Fuzhou Sunya Biotechnology Company.4 weeks later, we got parts one after another.
2017.08.06-2017.08.12
ALL of our parts were by Genscript Company, The Beijing Genomics Institute (BGI) Company and Fuzhou Sunya Biotechnology Company.4 weeks later, we got parts one after another.
2017.08.13-2017.08.19
ALL of our parts were by Genscript Company, The Beijing Genomics Institute (BGI) Company and Fuzhou Sunya Biotechnology Company.4 weeks later, we got parts one after another.
2017.08.20-2017.08.26
ALL of our parts were by Genscript Company, The Beijing Genomics Institute (BGI) Company and Fuzhou Sunya Biotechnology Company.4 weeks later, we got parts one after another.
2017.08.27-2017.09.02
We held CCiC
We planted wild-type tobacco and the southeast Sedum ,then we prepared the barrel, and began to prepare hydroponics experimental supplies. we Determination of Heavy Metals in Fertilizers.
2017.09.03-2017.09.09
We held CCiC
We planted wild-type tobacco and the southeast Sedum ,then we prepared the barrel, and began to prepare hydroponics experimental supplies. we Determination of Heavy Metals in Fertilizers.
2017.09.10-2017.09.16
We finished interlab work
After getting genes, we began to transform them to the Bacillus megaterium.And standardizing
2017.09.17-2017.09.23
We finished interlab work
After getting genes, we began to transform them to the Bacillus megaterium.And standardizing
2017.09.24-2017.09.30
In the two weeks, we carried on the induced expression and SDS-page.
2017.10.01-2017.10.07
After the Lpp-OmpA-MBP (Pb) and DsbA-MBP (Pb) were transferred to Bacillus megaterium, and cultured them in beef extract peptone medium with lead + xylose for 24 hours. Then we measured the growth curve.
We started preparing the experimental of Pb into wild-type tobacco
After GST-CRS5 was transferred to Bacillus megaterium, we measured the growth curve.
We started preparing the experimental of Cd into Sedum alfredii Hance
2017.10.08-2017.10.14
We finished the Formal experimental treatment of Pb into wild-type tobacco
2017.10.15-2017.10.21
We finished the Formal experimental treatment of Cd into Sedum alfredii Hance and take sample collection and treatment of plant, then analysis data.
