Team:Evry/Notebook/Interlabstudy/week8

From 2014.igem.org

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     <center>Figure 6: PCR mix Preparation for One Taq PCR colony  </center>
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     <center>Table 3: PCR mix Preparation for One Taq PCR colony  </center>
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49 µl of mix were distributed in each PCR tubes. The One Taq PCR program was applied.
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    <center>Figure 4:IGEM One Taq PCR program termocycling conditions </center>
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     <center>Figure 2: 1% agarose gel of PCR products from 08.12.2014 after PCR clean up. Lane 1 and 7: Purple 2-Log ladder NEB, Lane 2: BBa_J23101 purified PCR product, Lane 3: BBa_J23115 purified PCR product, Lane 4: BBa_E0240 purified PCR product, Lane 5: BBa_I20260 purified PCR product and Lane 6: pBHR1 purified PCR product  </center>
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     <center>Figure 2:   </center>
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Revision as of 12:55, 19 August 2014

Week 8

Interlab Study

08.18.2014

PCR colonies were performed on 8 colonies for each Biobrick (BBa_E0240 and BBa_I20260).

Sequencing results were received for BBa_J23101 (26DJ62 and 26DJ63) and BBa_J23115 (26DJ64 and 26DJ65) PCR products.
26DJ62

TCAGANNAAAAAAATCCTTAGCTNNCGCTAAGGATGANNTCTGGAATTCGCGGCCGCTTCTAGAG'''TTTACAGCTAGCTCAGTCCTAGGTATTATGCTAGC'''TACTAGTAG CGGCCGCTGCAGTCCGGCAAAAAAGGGCAAGGTGTCACCACCCTGCCCTTTTTCTTTAAAACCGAAAAGATTACTTCGCGTTATGCAGGCTTCCTCGCTCACTGACTC GCTGCGCTCGGTCGTTCGGCTGCGGCGAGCGGTATCAGCTCACTCAGAG


26DJ63

GAGCGCANCGAGTCAGTGAGCGAGGAAGCCTGCATAACGCGAAGTAATCTTTTCGGTTTTAAAGAAAAAGGGCAGGGTGGTGACACCTTGCCCTTTTTTGCCGGACTG CAGCGGCCGCTACTAGTA'''GCTAGCATAATACCTAGGACTGAGCTAGCTGTAAA'''CTCTAGAAGCGGCCGCGAATTCCAGAAATCATCCTTAGCGAAAGCTAAGGATTTTT TTTATCTGAAATTCTGCCTCGTGATACGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAGACGCAAGGTGGCAAA


26DJ64

TCAGANNAAAAAAATCCTTAGCTNNCGCTAAGGATGATTTCTGGAATTCGCGGCCGCTTCTAGAG'''GTTTATAGCTAGCTCA[[T]]CCT[[A]]GGTACAATGCTAGC'''TACTAG TAGCGGCCGCTGCAGTCCGGCAAAAAAGGGCAAGGTGTCACCACCCTGCCCTTTTTCTTTAAAACCGAAAAGATTACTTCGCGTTATGCAGGCTTCCTCGCTCACTGAC TCGCTGCGC TCGGTCGTTCGGCTGCGGCGAGCGGTATCAGCTCACTCAGG


26DJ65

CNGCGAGTCAGTNAGCGAGGAAGCCTGCANNACGCGAAGTNATCTTTTTCGGTTTTAAAGAAAAAGGGCAGGGTGGTGACACCTNGCCCTTTTTTGCCGGACTGCAGC GGCCGCTACTAGTA'''GCTAGCATTGTACCTAGGACTGAGCTAGCTATAAA'''CTCTAGAAGCGGCCGCGAATTCCAGAAATCATCCTTAGCGAAAGCTAAGGATTTTTTTTA TCTGAAATTCTGCCTCGTGATACGCCTATTTTTATAGGTTAATGTCATGATAATAATGGTTTCTTAAC

The mapping was perfect for BBa_J23101. There were 2 mismatches for BBa-J23115, one at 17 and the other t 21 on the promoter sequence that correspond to the K823012 as mentioned on 2014.igem.org/Tracks/Measurement/Measurement_Worksheet. The mapping was perfect between sequencing reads and K823012 sequence was perfect.
Colony PCRs were performed on 8 colonies for BBa_E0240 and BBa_I20260.
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Table 3: PCR mix Preparation for One Taq PCR colony
49 µl of mix were distributed in each PCR tubes. The One Taq PCR program was applied.
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Figure 4:IGEM One Taq PCR program termocycling conditions

08.19.2014

Preparation of a 1% agarose gel: 1.01 g of Top Vision agarose (Thermo Scientific) + 100 ml of TAE 1X. Microwave 30s by 30s until agarose total dissolution Gel was cooling down until to be lukewarm, one BET drop was added. Gel was loaded with 10µl per sample previously added with 2 µl of loading dye 6X, and 5 µl for ladders. Gel running 45 minutes at 100 mV in TAE 1X buffer.
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Figure 2:
We decided to amplify the colony 1 for BBa_E0240 and BBa_I20260. So PCR products were purified via with the GeneJET purification kit (Thermo Scientific)followed by DNA quantification with the NanoDrop 2000 (Thermo Scientific). BBa_E0240 purified PCR product: 61 ng/µl and BBa_I20260 purified PCR product: 37.8 ng/µl.
Purified PCR product aliquot was send to sequencing. The sequencing number were 26DJ68, 69, 70, 71.

08.20.2014

08.21.2014

08.22.2014

08.23.2014

08.24.2014

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