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From 2011.igem.org

Wednesday

A few mistakes from last week, a fully booked PCR machine and a crowded freezer led to a morning of low spirits and around 12 O'clock it seemed there were nothing more that could be done. But after a lunch everything looked much brighter and positive thinking and troubleshooting kept us busy from then on - the A1 was ligated into the standard plasmid AND the expression vector. Around 2 O'clock another miracle happened as a missing BioBrick disappeared in the freezer and made sure that we had something to do for the rest of the day as well.

A1 The only one without any problems. So we ligated it with both the standard plasmid and a expression vector.

• A1+pSB1C3

• • Ligation

• • Transformation

• A1+ekspression vector

• • Double Digestion with Xba1 and Pst1

• • Ligation with the expression vector
    • Size of BioBrick J04500 = 220bp+3189=3409 bp
    • Size of BioBrick A1 = 1600bp
    • 3409/1600= ca. 2

• • Transformation

• Expressionvector'

• • Double Digestion with Pst1 and Spe1

A2

We can't find any DNA in our tubes and it seems that the PCR prefix and suffix addition went wrong. So we did that again.

• Addition of prefix and suffix with PCR

B1

After a long and hard search after the digested B1 it was finally found and ligated with the standard plasmid.

• Found the B1 biobrick
• Ligation with pSB1C3
• Transformation of XL1-Blue