LOCUS       pENTR2b-lifeAct-        3060 bp DNA     circular SYN 11-JUN-2020
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 3060)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 7, 2024 from SnapGene Viewer 7.2.1
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..3060
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     terminator      103..189
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      281..308
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     protein_bind    358..457
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    458..493
                     /label=MCS 1
                     /note="multiple cloning site, part 1"
     misc_feature    494
                     /label=overhang+Kpn1
     misc_feature    495
                     /label=T changed to G for Kozacs seq restauration
     CDS             501..551
                     /codon_start=1
                     /label=LifeAct
                     /translation="MGVADLIKKFESISKEE"
     CDS             573..1268
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     misc_feature    1269..1271
                     /label=stop codon
     misc_feature    1272..1274
                     /label=2nd stop codon
                     /note="Inserted by me. "
     misc_feature    1275
                     /label=XhoI+overhang
     misc_feature    1276..1285
                     /label=MCS 2
                     /note="multiple cloning site, part 2"
     protein_bind    complement(1289..1388)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             1511..2320
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      2410..2998
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgctagca tggatctcgg
       61 ggacgtctaa ctactaagcg agagtaggga actgccaggc atcaaataaa acgaaaggct
      121 cagtcggaag actgggcctt tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt
      181 aggacaaatc cgccgggagc ggatttgaac gttgtgaagc aacggcccgg agggtggcgg
      241 gcaggacgcc cgccataaac tgccaggcat caaactaagc agaaggccat cctgacggat
      301 ggcctttttg cgtttctaca aactcttcct gttagttagt tacttaagct cgggccccaa
      361 ataatgattt tattttgact gatagtgacc tgttcgttgc aacaaattga taagcaatgc
      421 ttttttataa tgccaacttt gtacaaaaaa gcaggctggc gccggaacca attcagtcga
      481 ctggatccgg taccgccacc atgggcgtgg ccgacctgat caagaagttc gagagcatca
      541 gcaaggagga gggggatcca ccggtcgcca ccatggtgag caagggcgag gcagtgatca
      601 aggagttcat gcggttcaag gtgcacatgg agggctccat gaacggccac gagttcgaga
      661 tcgagggcga gggcgagggc cgcccctacg agggcaccca gaccgccaag ctgaaggtga
      721 ccaagggtgg ccccctgccc ttctcctggg acatcctgtc ccctcagttc atgtacggct
      781 ccagggcctt catcaagcac cccgccgaca tccccgacta ctataagcag tccttccccg
      841 agggcttcaa gtgggagcgc gtgatgaact tcgaggacgg cggcgccgtg accgtgaccc
      901 aggacacctc cctggaggac ggcaccctga tctacaaggt gaagctccgc ggcaccaact
      961 tccctcctga cggccccgta atgcagaaga agacaatggg ctgggaagcg tccaccgagc
     1021 ggttgtaccc cgaggacggc gtgctgaagg gcgacattaa gatggccctg cgcctgaagg
     1081 acggcggccg ctacctggcg gacttcaaga ccacctacaa ggccaagaag cccgtgcaga
     1141 tgcccggcgc ctacaacgtc gaccgcaagt tggacatcac ctcccacaac gaggactaca
     1201 ccgtggtgga acagtacgaa cgctccgagg gccgccactc caccggcggc atggacgagc
     1261 tgtacaagta atagctcgag atatctagac ccagctttct tgtacaaagt tggcattata
     1321 agaaagcatt gcttatcaat ttgttgcaac gaacaggtca ctatcagtca aaataaaatc
     1381 attatttgcc atccagctgc agctctggcc cgtgtctcaa aatctctgat gttacattgc
     1441 acaagataaa aatatatcat catgaacaat aaaactgtct gcttacataa acagtaatac
     1501 aaggggtgtt atgagccata ttcaacggga aacgtcgagg ccgcgattaa attccaacat
     1561 ggatgctgat ttatatgggt ataaatgggc tcgcgataat gtcgggcaat caggtgcgac
     1621 aatctatcgc ttgtatggga agcccgatgc gccagagttg tttctgaaac atggcaaagg
     1681 tagcgttgcc aatgatgtta cagatgagat ggtcagacta aactggctga cggaatttat
     1741 gcctcttccg accatcaagc attttatccg tactcctgat gatgcatggt tactcaccac
     1801 tgcgatcccc ggaaaaacag cattccaggt attagaagaa tatcctgatt caggtgaaaa
     1861 tattgttgat gcgctggcag tgttcctgcg ccggttgcat tcgattcctg tttgtaattg
     1921 tccttttaac agcgatcgcg tatttcgtct cgctcaggcg caatcacgaa tgaataacgg
     1981 tttggttgat gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg aacaagtctg
     2041 gaaagaaatg cataaacttt tgccattctc accggattca gtcgtcactc atggtgattt
     2101 ctcacttgat aaccttattt ttgacgaggg gaaattaata ggttgtattg atgttggacg
     2161 agtcggaatc gcagaccgat accaggatct tgccatccta tggaactgcc tcggtgagtt
     2221 ttctccttca ttacagaaac ggctttttca aaaatatggt attgataatc ctgatatgaa
     2281 taaattgcag tttcatttga tgctcgatga gtttttctaa tcagaattgg ttaattggtt
     2341 gtaacattat tcagattggg ccccgttcca ctgagcgtca gaccccgtag aaaagatcaa
     2401 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     2461 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     2521 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
     2581 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     2641 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     2701 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     2761 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     2821 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     2881 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     2941 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     3001 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
//
