LOCUS       Exported                6161 bp DNA     circular SYN 28-NOV-2022
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6161)
  AUTHORS   sara
  TITLE     Direct Submission
  JOURNAL   Exported Nov 28, 2022 from SnapGene 5.3.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6161
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        2..20
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     5'UTR           45..87
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     regulatory      110..119
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             116..169
                     /codon_start=1
                     /label=start of CDS
                     /translation="MASPKKKRKVEASGGLQN"
     CDS             125..145
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             170..880
                     /codon_start=1
                     /gene="mgfp5"
                     /product="GFP with folding enhancement mutations"
                     /label=mgfp5
                     /note="suitable for expression in plants due to removal of 
                     a cryptic intron"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFNDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKAN
                     FKTRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     misc_feature    887..3034
                     /label=BTGH84 (-20 aa of signal peptide)
     misc_feature    1610..1612
                     /label=D242 in BtGH84 structure by Dennis et al.
                     /note="Dennis, R. J., Taylor, E. J., MacAuley, M. S., 
                     Stubbs, K. A., Turkenburg, J. P., Hart, S. J., … Davies, G.
                     J. (2006). Structure and mechanism of a bacterial 
                     beta-glucosaminidase having O-GlcNAcase activity. Nature 
                     Structural and Molecular Biology, 13(4), 365–371. 
                     https://doi.org/10.1038/nsmb1079"
     misc_feature    1610..1612
                     /label=D242A (""dead"" BTGH mutation)
     CDS             3044..3064
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3068..3088
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKA"
     CDS             3092..3112
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     3'UTR           3135..3260
                     /label=Xenopus globin 3'-UTR
                     /note="3'-UTR of the major beta-globin gene of Xenopus 
                     laevis"
     promoter        complement(3349..3367)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3374..3390)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      complement(3532..3987)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        4016..4120
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4121..4981
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      5152..5740
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    6028..6049
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        6064..6094
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6102..6118
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     6126..6142
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 aaattaaccc tcactaaagg gaacaaaagc tggagctaag cttgcttgtt ctttttgcag
       61 aagctcagaa taaacgctca actttggcag atctgaattc ctgcagcccg ccaccatggc
      121 ctctcctaag aaaaagagga aagtggaagc atcaggcggg ctgcagaata gtaaaggaga
      181 agaacttttc actggagttg tcccaattct tgttgaatta gatggtgatg ttaatgggca
      241 caaattttct gtcagtggag agggtgaagg tgatgcaaca tacggaaaac ttacccttaa
      301 atttatttgc actactggaa aactacctgt tccatggcca acacttgtca ctactttcac
      361 ttatggtgtt caatgctttt caagataccc agatcatatg aagcggcacg acttcttcaa
      421 gagcgccatg cctgagggat acgtgcagga gaggaccatc ttcttcaatg acgacgggaa
      481 ctacaagaca cgtgctgaag tcaagtttga gggagacacc ctcgtcaaca ggatcgagct
      541 taagggaatc gatttcaagg aggacggaaa catcctcggc cacaagttgg aatacaacta
      601 caactcccac aacgtataca tcatggccga caagcaaaag aacggcatca aagccaactt
      661 caagacccgc cacaacatcg aagacggcgg cgtgcaactc gctgatcatt atcaacaaaa
      721 tactccaatt ggcgatggcc ctgtcctttt accagacaac cattacctgt ccacacaatc
      781 tgccctttcg aaagatccca acgaaaagag agaccacatg gtccttcttg agtttgtaac
      841 agctgctggg attacacatg gcatggatga actatacaaa ggcggacaga atgtttctct
      901 ccaacctcca cctcagcagc tgatagtgca gaacaagact attgacctcc cggcagtcta
      961 ccagttgaat ggaggcgagg aggctaaccc ccacgctgtc aaggtactta aggaattgtt
     1021 gtccggcaag cagagctcaa aaaaggggat gctgattagt atcggggaga aaggagataa
     1081 atcagtaaga aagtacagta ggcagatacc agaccacaaa gaaggttact acctctctgt
     1141 taacgagaag gagatagtgc tggcaggcaa cgacgagcgc ggaacgtatt acgccttgca
     1201 aacttttgct cagctcttga aggacgggaa gttgcccgag gtagagataa aagactaccc
     1261 cagtgtccgg tataggggcg tggtagaggg gttctacggc actccatggt ctcatcaggc
     1321 tagactgagt cagttgaagt tctatggcaa aaacaagatg aacacctata tatatggtcc
     1381 caaggacgac ccgtatcact ccgctccaaa ctggcggctc ccctatcctg acaaggaggc
     1441 tgctcagctg caggagcttg tggctgttgc taacgaaaac gaggtcgatt ttgtatgggc
     1501 aattcatccc gggcaagata tcaagtggaa caaggaggat cgggatctgc tcctggccaa
     1561 gttcgaaaag atgtatcaac tgggggtccg atctttcgcc gtgttctttg ccgatatcag
     1621 cggggaagga actaaccctc agaagcaggc agagttgctt aactatatcg atgagaagtt
     1681 tgcacaggtg aagcctgata ttaaccagct cgtgatgtgt cccacagagt ataacaagag
     1741 ctggagtaac cccaatggaa actacctcac tacacttggt gacaagctga acccctctat
     1801 ccagatcatg tggacagggg accgggttat atctgacata accagggacg gaatctcttg
     1861 gattaatgaa aggatcaagc gaccagccta catctggtgg aacttcccag tatctgatta
     1921 tgtcagagac cacctgctgc tggggcctgt gtacggcaat gatactacga tcgccaagga
     1981 aatgtccggc tttgtcacaa atcccatgga acacgctgag tcctctaaaa ttgctatcta
     2041 cagcgtggca tcatatgcct ggaatcccgc caagtatgac acctggcaga cttggaagga
     2101 tgcaatccgc acgatcctgc caagtgctgc agaggagctc gagtgtttcg ccatgcataa
     2161 cagcgacctg gggcccaatg gacatgggta tagacgagaa gaatcaatgg atatccagcc
     2221 tgccgcagag cggtttctga aagctttcaa agaaggaaaa aattacgaca aagccgattt
     2281 tgaaaccttg cagtacacct tcgagagaat gaaggagtct gccgacatcc tgctgatgaa
     2341 tacggaaaat aagccactga ttgtggagat cacaccttgg gtccaccaat tcaagttgac
     2401 cgctgaaatg ggcgaagagg ttctcaaaat ggtagagggg agaaacgaat cctacttctt
     2461 gcgcaagtac aaccatgtga aagctctcca acagcagatg ttctatattg accagacgtc
     2521 taaccagaac ccttatcaac caggtgtcaa gaccgccaca cgggtgataa agccactgat
     2581 agatcgcaca tttgcaacag ttgtgaaatt ttttaaccag aagttcaatg cccacctgga
     2641 cgccaccact gattacatgc cccacaagat gatcagcaac gtcgagcaaa ttaagaatct
     2701 ccccctgcag gtgaaagcaa atagggtcct gatctccccc gctaatgagg tggtcaagtg
     2761 ggccgctggt aattccgtgg aaatcgaact tgatgcaatc tatcctggtg agaatataca
     2821 gataaatttc ggcaaggacg caccctgcac ttggggccgc cttgagatca gcactgatgg
     2881 gaaagagtgg aaaaccgtgg acctgaagca gaaagagtcc cgcctttccg ctggactgca
     2941 gaaggctcca gtaaaattcg tccgattcac caacgtcagc gatgaggagc agcaggtcta
     3001 tttgcggcaa ttcgtcttga caattgagaa aaagactgga gaccccaaaa agaagcgcaa
     3061 agttgatcca aaaaaaaaaa ggaaagccga tcctaagaag aaacggaaag tgtgagcgga
     3121 tctggttacc actaaaccag cctcaagaac acccgaatgg agtctctaag ctacataata
     3181 ccaacttaca ctttacaaaa tgttgtcccc caaaatgtag ccattcgtat ctgctcctaa
     3241 taaaaagaaa gtttcttcac attctaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaacc
     3301 cccccccccc cccccctgca gggccaagtc ggccggtacc caattcgccc tatagtgagt
     3361 cgtattacaa ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta
     3421 cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg
     3481 cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaaggg caaattgtaa
     3541 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     3601 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     3661 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     3721 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     3781 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     3841 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     3901 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     3961 cgcttaatgc gccgctacag ggcgcgtccc aggtggcact tttcggggaa atgtgcgcgg
     4021 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     4081 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     4141 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     4201 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     4261 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     4321 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     4381 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     4441 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     4501 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     4561 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     4621 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     4681 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     4741 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     4801 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     4861 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     4921 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     4981 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     5041 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     5101 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     5161 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     5221 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     5281 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     5341 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     5401 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     5461 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     5521 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     5581 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     5641 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     5701 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     5761 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     5821 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     5881 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc
     5941 gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg
     6001 gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca
     6061 ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt
     6121 tcacacagga aacagctatg accatgatta cgccaagctc g
//
