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From 2011.igem.org

Contents 1 5. Labday 2 Miniprep 3 3A-assembly: Digestion 4 Protein modelling

5. Labday

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Miniprep

Investigators: Jakob, Julia

Comments: Trafo was performed on 8th of june and plates were incubated over night at 37°C. Clones were picked to do a miniprep.

Miniprep of parts from 8th of june and DNA concentration was measured with NanoDrop 2000c.

3A-assembly: Digestion

Investigators: Sophie, Theo, Sandra

Digestion of RBS and promotor to create a promotor-RBS library.

	sample	name
Promotor	BBa_J23104	S1
Promotor	BBa_J23110	S2
Promotor	BBa_J23116	S3
RBS	BBa_B0034	S4
RBS	BBa_B0032	S5
RBS	BBa_B0031	S6
Vector	pSB1C3	S7

Enzymes used for digestion:

	enzyme 1	enzyme 2
promotor (S1-S3)	EcoRI	SpeI
RBS (S4-S6)	Xbal	PstI
Vector (S7)	EcoRI	PstI

Protein modelling

Investigators: Rüdiger

3cvr bacterial ligase

FSELQLNRLNLSSLPDNLPPQ

ITVLEITQNALISLPELPAS

LEYLDACDNRLSTLPELPAS

LKHLDVDNNQLTXLPELPAL

LEYINADNNQLTXLPELPTS

LEVLSVRNNQLTFLPELPES

EIFFRCRENRITHIPENILSLDP

Extracted ideal consensus, by consensus sequence and 3d rational analysis:

LEVLDVSNNQLTSLPDNLPAS

N-Terminal Hagfish 2z66 CPSRCSCSGTEIRCNSKGLTSVPTGIPSSATRLELESNKLQSLPHGVFDK LTQLTK Consensus: xLxxLxxLxL C-Terminal Hagfish 2z62 LKELALDTNQLKSVPDGIFDR LTSLQKIWLHTNPWDCSCPRIDY LSRWLNKNSQKEQGSAKCSGSGKPVRSIICP LxxLxLxxNxLx Consensus

Sequences 1-6 sent to I-Tasser for 3D structure prediction: Rational1: LxALHCSxNxLxSLPxxLPxx LxHLACSxNxLxSLPxxLPxx

CPSRCSCSGTEIRCNSKGLTSVPTGIPSS ATRLELESNKLQSLPHGVFDK LTALHKSNNQLTSLPDNLPAS LEHLAVSNNQLTSLPDNLPAS LEALHVSNNQLTSLPDNLPAS LEHLAVSNNQLTSLPDNLPAS LEALHVSNNQLTSLPDNLPAS LEHLAVSNNQLTSLPDNLPAS LKELALDTNQLKSVPDGIFDR LTSLQKIWLHTNPWDCSCPRIDY LSRWLNKNSQKEQGSAKCSGSGKPVRSIICP

Rational2: LxxLxCSxNHLHSLPxxLPxx

CPSRCSCSGTEIRCNSKGLTSVPTGIPSS ATRLELESNKLQSLPHGVFDK LTQLTKSNNHLHSLPDNLPAS LEVLDVSNNHLHSLPDNLPAS LEVLDVSNNHLHSLPDNLPAS LEVLDVSNNHLHSLPDNLPAS LEVLDVSNNHLHSLPDNLPAS LEVLDVSNNHLHSLPDNLPAS LKELALDTNQLKSVPDGIFDR LTSLQKIWLHTNPWDCSCPRIDY LSRWLNKNSQKEQGSAKCSGSGKPVRSIICP

Rational3: LxxLxCSxNxLxSLPHHLPxx

CPSRCSCSGTEIRCNSKGLTSVPTGIPSS ATRLELESNKLQSLPHGVFDK LTQLTKSNNQLTSLPHHLPAS LEVLDVSNNQLTSLPHHLPAS LEVLDVSNNQLTSLPHHLPAS LEVLDVSNNQLTSLPHHLPAS LEVLDVSNNQLTSLPHHLPAS LEVLDVSNNQLTSLPHHLPAS LKELALDTNQLKSVPDGIFDR LTSLQKIWLHTNPWDCSCPRIDY LSRWLNKNSQKEQGSAKCSGSGKPVRSIICP

Rational4: LxxLxCSxNxLxSLPxHLPHx

CPSRCSCSGTEIRCNSKGLTSVPTGIPSS ATRLELESNKLQSLPHGVFDK LTQLTKSNNQLTSLPDHLPHS LEVLDVSNNQLTSLPDHLPHS LEVLDVSNNQLTSLPDHLPHS LEVLDVSNNQLTSLPDHLPHS LEVLDVSNNQLTSLPDHLPHS LEVLDVSNNQLTSLPDHLPHS LKELALDTNQLKSVPDGIFDR LTSLQKIWLHTNPWDCSCPRIDY LSRWLNKNSQKEQGSAKCSGSGKPVRSIICP

Rational5: LHxLxCSxNxLxSLPxxLPxH LHVLDVSNNQLTSLPDNLPAH

CPSRCSCSGTEIRCNSKGLTSVPTGIPSS ATRLELESNKLQSLPHGVFDK LHQLTKSNNQLTSLPDNLPAH LHVLDVSNNQLTSLPDNLPAH LHVLDVSNNQLTSLPDNLPAH LHVLDVSNNQLTSLPDNLPAH LHVLDVSNNQLTSLPDNLPAH LHVLDVSNNQLTSLPDNLPAH LKELALDTNQLKSVPDGIFDR LTSLQKIWLHTNPWDCSCPRIDY LSRWLNKNSQKEQGSAKCSGSGKPVRSIICP

Rational6: LHHLxCSxNxLxSLPxxLPxx LHHLDVSNNQLTSLPDNLPAS

CPSRCSCSGTEIRCNSKGLTSVPTGIPSS ATRLELESNKLQSLPHGVFDK LHHLTKSNNQLTSLPDNLPAS LHHLDVSNNQLTSLPDNLPAS LHHLDVSNNQLTSLPDNLPAS LHHLDVSNNQLTSLPDNLPAS LHHLDVSNNQLTSLPDNLPAS LHHLDVSNNQLTSLPDNLPAS LHHLDVSNNQLTSLPDNLPAS LKELALDTNQLKSVPDGIFDR LTSLQKIWLHTNPWDCSCPRIDY LSRWLNKNSQKEQGSAKCSGSGKPVRSIICP