BBa_K2022012 1 BBa_K2022012 INPNC-GFP, i.e. membrane-anchored GFP 2016-10-11T11:00:00Z 2016-10-12T03:35:40Z To make this part, we combined BBa_K811005 (INPNC) with BBa_E0040 (GFP). First, we digested BBa_K811005 with BamHI and PstI. Then, we PCR amplified BBa_E0040 using Suffix-R2, and a forward oligonucleotides that replaced the prefix with a BamHI site (aggcggatccggtatgcgtaaaggagaagaacttttcactg). This PCR product was then also digested with BamHI and PstI. These two products were then ligated together, and transformed into E. coli host (DH5-alpha derivative) A commonly used reporter is Green Fluorescent Protein (GFP) (BBa_E0040). Here, we have fused it to the C-terminus of the Ice Nucleation Protein, N- and C-termini (BBa_K811005), which is a well-characterised membrane anchor. As such, this part is intended to function like GFP, but simply localised to the membrane of cells false false _2489_ 31054 31054 9 false Pennsylvania (2012) designed their INPNC part with the BamHI site, to make it readily fusible to other proteins; we thank them for their contribution in making the assembly and design processes as simple as possible. As such, the only design consideration was to use the oligonucleotide aggcggatccggtatgcgtaaaggagaagaacttttcactg to add a BamHI site upstream of the GFP open reading frame false Jessica Mazalo, Daniel Winter annotation2493571 1 double stop range2493571 1 1675 1680 annotation2493566 1 start range2493566 1 1 3 annotation2493572 1 INPNC range2493572 1 1 957 annotation2493573 1 GFP range2493573 1 961 1680 BBa_K2022012_sequence 1 atgaccctggacaaagctctggttctgcgtacctgcgctaacaacatggctgaccactgcggtctgatctggccggcttctggtaccgttgaatctcgttactggcagtctacccgtcgtcacgaaaacggtctggttggtctgctgtggggtgctggtacctctgctttcctgtctgttcacgctgacgctcgttggatcgtttgcgaagttgctgttgctgacatcatctctctggaagaaccgggtatggttaaattcccgcgtgctgaagttgttcacgttggtgaccgtatctctgcttctcacttcatctctgctcgtcaggctgacccggcttctacctctacctctacctctacctctaccctgaccccgatgccgaccgctatcccgaccccgatgccggctgttgcttctgttaccctgccggttgctgaacaggctcgtcacgaagttttcgacgttgcttctgtttctgctgctgctgctccggttaacaccctgccggttaccaccccgcagaacctgcaaaccgctacctacggttctaccctgtctggtgacaaccactctcgtctgatcgctggttacggttctaacgaaaccgctggtaaccactctgacctgatcggtggtcacgactgcaccctgatggctggtgaccagtctcgtctgaccgctggtaaaaactctgttctgaccgctggtgctcgttctaaactgatcggttctgaaggttctaccctgtctgctggtgaagactctaccctgatcttccgtctgtgggacggtaaacgttaccgtcagctggttgctcgtaccggtgaaaacggtgttgaagctgacatcccgtactacgttaacgaagacgacgacatcgttgacaaaccggacgaagacgacgactggatcgaagttaaaggtggcggtagtggcggtgcaggaggcggatccggtatgcgtaaaggagaagaacttttcactggagttgtcccaattcttgttgaattagatggtgatgttaatgggcacaaattttctgtcagtggagagggtgaaggtgatgcaacatacggaaaacttacccttaaatttatttgcactactggaaaactacctgttccatggccaacacttgtcactactttcggttatggtgttcaatgctttgcgagatacccagatcatatgaaacagcatgactttttcaagagtgccatgcccgaaggttatgtacaggaaagaactatatttttcaaagatgacgggaactacaagacacgtgctgaagtcaagtttgaaggtgatacccttgttaatagaatcgagttaaaaggtattgattttaaagaagatggaaacattcttggacacaaattggaatacaactataactcacacaatgtatacatcatggcagacaaacaaaagaatggaatcaaagttaacttcaaaattagacacaacattgaagatggaagcgttcaactagcagaccattatcaacaaaatactccaattggcgatggccctgtccttttaccagacaaccattacctgtccacacaatctgccctttcgaaagatcccaacgaaaagagagaccacatggtccttcttgagtttgtaacagctgctgggattacacatggcatggatgaactatacaaataataa igem2sbol 1 iGEM to SBOL conversion Conversion of the iGEM parts registry to SBOL2.1 Chris J. Myers James Alastair McLaughlin 2017-03-06T15:00:00.000Z