LOCUS       Exported                7203 bp ds-DNA     circular SYN 15-FEB-2021
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7203)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Dec 20, 2021 from SnapGene Viewer 5.1.5
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7203
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      complement(3..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: RIGHT"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     misc_feature    653..814
                     /label=minitol2-L
                     /note="color: #ff0000"
     protein_bind    820..840
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
                     /note="color: #31849b; direction: RIGHT"
     misc_feature    841..856
                     /label=QUAS
                     /note="color: #a6acb3"
     misc_feature    859..874
                     /label=QUAS#
                     /note="color: #ff0000"
     misc_feature    879..894
                     /label=QUAS
                     /note="color: #a6acb3"
     misc_feature    899..914
                     /label=QUAS
                     /note="color: #a6acb3"
     misc_feature    917..932
                     /label=QUAS
                     /note="color: #a6acb3"
     misc_feature    974..1006
                     /label=minimal promoter
                     /note="color: #a6acb3"
     primer_bind     1016..1030
                     /label=T4 Mlu1 zsG-p2A del F
                     /note="color: black; sequence: GACCTCGAAGACGCG; added: 
                     2019-07-15"
     protein_bind    1031..1055
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
                     /note="color: #31849b; direction: RIGHT"
     CDS             1062..1085
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /note="This feature has 2 segments:
                        1: 1062 .. 1070 / #cc99b2
                        2: 1071 .. 1085 / #cc99b2 / enterokinase site
                     Cleavage site after base 1085"
                     /translation="DLYDDDDK"
     CDS             1086..1775
                     /codon_start=1
                     /product="Zoanthus green fluorescent protein"
                     /label=ZsGreen1
                     /note="mammalian codon-optimized"
                     /note="color: #05fd14"
                     /translation="AQSKHGLTKEMTMKYRMEGCVDGHKFVITGEGIGYPFKGKQAINL
                     CVVEGGPLPFAEDILSAAFMYGNRVFTEYPQDIVDYFKNSCPAGYTWDRSFLFEDGAVC
                     ICNADITVSVEENCMYHESKFYGVNFPADGPVMKKMTDNWEPSCEKIIPVPKQGILKGD
                     VSMYLLLKDGGRLRCQFDTVYKAKSVPRKMPDWHFIQHKLTREDRSDAKNQKWHLTEHA
                     IASGSALP"
     primer_bind     1776..1795
                     /label=T4 EcoR1 Numb-p2A F
                     /note="color: black; sequence: 
                     TAGAACTTGCATGCGGAAGCGGAGCTAC"
     CDS             1791..1847
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /note="color: #cc99b2
                     Cleavage site after base 1844"
                     /translation="ATNFSLLKQAGDVEENPGP"
     primer_bind     complement(1828..1847)
                     /label=T4 p2A-Flag R
                     /note="color: black; sequence: 
                     ttgtagtcAGGTCCAGGGTTCTCCTCCA"
     primer_bind     complement(1848..1862)
                     /label=T4 Bgl2 zsG-p2A del R
                     /note="color: black; sequence: tcgacCTCGAGatct; added: 
                     2019-07-15"
     primer_bind     1854..1870
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: RIGHT"
     misc_feature    1910..3114
                     /label=Ocean pout Terminator
                     /note="color: #ff0000"
     misc_feature    3123..3830
                     /label=Crystallin Alpha Promoter
                     /note="color: #ff00ff"
     primer_bind     3808..3848
                     /label=T4 Age1 a cry - mCherry F
                     /note="color: black; sequence: 
                     ctgtGGATCCACCGGTCGCCACCATGGTGAGCAAGGGCGAG"
     CDS             3831..4541
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /note="color: #ff0000"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(4526..4562)
                     /label=T4 Fse1 mCherry-KY64 R
                     /note="color: black; sequence: 
                     TGATCTAGAGTCgcggccgctTTACTTGTACAGCTCG"
     polyA_signal    4664..4745
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     misc_feature    4795..5002
                     /label=minitol2-R
                     /note="color: #ff0000"
     promoter        complement(5015..5033)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     primer_bind     complement(5054..5070)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     protein_bind    5078..5094
                     /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)."
                     /note="color: #31849b"
     promoter        complement(5102..5132)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This reverse directional feature has 3 segments:
                        1: 5102 .. 5108 / #ffffff / -10
                        2: 5109 .. 5126 / #ffffff
                        3: 5127 .. 5132 / #ffffff / -35"
     protein_bind    5147..5168
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
                     /note="color: #31849b"
     rep_origin      complement(5456..6044)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     CDS             complement(6215..7075)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 6215 .. 7006 / #ccffcc
                        2: 7007 .. 7075 / #ccffcc / signal sequence
                     Cleavage site after base 7006"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(7076..7180)
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: LEFT"
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tgggtaccca
      661 gaggtgtaaa aagtactcaa aaattttact caagtgaaag tacaagtact tagggaaaat
      721 tttactcaat taaaagtaaa agtatctggc tagaatctta cttgagtaaa agtaaaaaag
      781 tactccatta aaattgtact tgagtattct cgaggtcgac aactttgtat agaaaagttg
      841 gggtaatcgc ttatcctcgg ataaacaatt atcctcacgg gtaatcgctt atccgctcgg
      901 gtaatcgctt atcctcgggt aatcgcttat ccttgcaagg gtcgactcta gagggtatat
      961 aatggatccc atcgcgtctc agcctcactt tgagctcctc cacacgaatt ccctcgacct
     1021 cgaagacgcg tcaagtttgt acaaaaaagc aggtcaccat ggatctgtac gacgatgacg
     1081 ataaggccca gtccaagcac ggcctgacca aggagatgac catgaagtac cgcatggagg
     1141 gctgcgtgga cggccacaag ttcgtgatca ccggcgaggg catcggctac cccttcaagg
     1201 gcaagcaggc catcaacctg tgcgtggtgg agggcggccc cttgcccttc gccgaggaca
     1261 tcttgtccgc cgccttcatg tacggcaacc gcgtgttcac cgagtacccc caggacatcg
     1321 tcgactactt caagaactcc tgccccgccg gctacacctg ggaccgctcc ttcctgttcg
     1381 aggacggcgc cgtgtgcatc tgcaacgccg acatcaccgt gagcgtggag gagaactgca
     1441 tgtaccacga gtccaagttc tacggcgtga acttccccgc cgacggcccc gtgatgaaga
     1501 agatgaccga caactgggag ccctcctgcg agaagatcat ccccgtgccc aagcagggca
     1561 tcttgaaggg cgacgtgagc atgtacctgc tgctgaagga cggtggccgc ttgcgctgcc
     1621 agttcgacac cgtgtacaag gccaagtccg tgccccgcaa gatgcccgac tggcacttca
     1681 tccagcacaa gctgacccgc gaggaccgca gcgacgccaa gaaccagaag tggcacctga
     1741 ccgagcacgc catcgcctcc ggctccgcct tgcccgcatg cggaagcgga gctactaact
     1801 tcagcctgct gaagcaggct ggagacgtgg aggagaaccc tggacctaga tctcgaggtc
     1861 gacggtatcg ataagcttga tatcgaattc ctgcagcccg ggggatccac tagtaattaa
     1921 gtctcagcca ccgttaactg aacatgtcaa aacctgtgga gactgttgag atttgatgtt
     1981 ctgaaaagat aaagcctata aataaaatgt tgcccaaatt tcctgcctga tgtttttctt
     2041 tgtctttgct acatggcttt gctgctcgga tcggctcact ctgtgtatgc cacgttcact
     2101 ttgtactctc cttctcacgg taggtttatt atttttagat gtgcagttag tttctgtgaa
     2161 ataacacacc acacactgat attgtctgtg cattgacttg gtgagtgcac attgtttttg
     2221 atcttgacat atttatattt gattgatcag gtgaactgtg tgaatctaaa gtgctccata
     2281 cagatgttct gcattgaaaa tattctcatt ttattagtgg aagtgagtgt atgccacatc
     2341 caatcaattt cagcaaacac cccagtatga tttaatgcaa aaaaatgaag gtatcaaaca
     2401 cgcattacta ctttgcagtt aaatatttaa catttattcc aacacgaaaa aaagcagtaa
     2461 ataacacttt gacaaacacg tcaggacatc ttatttttgt caccctcaca ggcaatttag
     2521 tataatatat tatatatata tatatatcat ataataatat tcagtataat atatatatat
     2581 atatcatatt ataatattca gtataatata aaacacaaac acatatatgt ataatataat
     2641 ataacatttt tatttattga gatgcctcta tggaccgtgt tataagaagt aaagatcagg
     2701 agaagtaaac atgaagtgta attatgaata ctgatgttaa attaagctat gatgagtttt
     2761 cactgttaat ttaccatctc aattaaatgt tgatgcctcc atgaccaagt taagcagatg
     2821 agactgagac aactgtagaa gacaagatgt tcactttgct gaatatagct ggcttgacag
     2881 ttatctatga ctctataaat atatatatat tttttttttt ataaaatgat ttatttataa
     2941 ctatatatcc atttctcaga caggtgcttc atatccctca ctcccgtagc tgtccatgct
     3001 ggatctgtcc ccgttgtttt taaaaagcta aataagttat taacatgact gcatccagcg
     3061 agccaaacct gtctggtgta cagctaccag agaagcttga gatcctagtc accggcggcc
     3121 gctaatatgg caataaacga tcttcagaga aacttgcatt ggtaaatagt ttcagcctat
     3181 gcatatgtag ttattcaaac tgtaatgact atcttacagt attatatgtt tataatagtt
     3241 tgatccaata aatccagatt tatatgcttt tcaacataaa attgtgaaat gcacagaatg
     3301 tacatataca atacatttcc tacaataatg ctctcttttc tattaatagt gtgcattcag
     3361 tgcaggatgt tatgtattta ttttctgtag tgcaaacttt ttcaaagaat tttccagtct
     3421 aagcttttca gtcaagaaaa aaaaaagcga ttgattatct gtgatattta aaaaaattag
     3481 ctcttaaatc tggtgaacca agtggccaca acacattaaa tactgacgtt ctacagccat
     3541 gtagcttcat ctggtctggt ttgttttggc aggcacttat tttccggtga tagttggcca
     3601 caagagtatt gtctgagcca ttcagtgcta gactgtcatt ctcaggtcag agtccacccc
     3661 gctgattctg ctgacaacac tgttcccacc atgagataat gccattccag agagatccat
     3721 ttgtaagccc ctctttctgc agcacaggta tataaccagg ggtctgcctc cactaaggcc
     3781 ggcacacatc atttggggat ctttgtactg tggatccacc ggtcgccacc atggtgagca
     3841 agggcgagga ggataacatg gccatcatca aggagttcat gcgcttcaag gtgcacatgg
     3901 agggctccgt gaacggccac gagttcgaga tcgagggcga gggcgagggc cgcccctacg
     3961 agggcaccca gaccgccaag ctgaaggtga ccaagggtgg ccccctgccc ttcgcctggg
     4021 acatcctgtc ccctcagttc atgtacggct ccaaggccta cgtgaagcac cccgccgaca
     4081 tccccgacta cttgaagctg tccttccccg agggcttcaa gtgggagcgc gtgatgaact
     4141 tcgaggacgg cggcgtggtg accgtgaccc aggactcctc cctgcaggac ggcgagttca
     4201 tctacaaggt gaagctgcgc ggcaccaact tcccctccga cggccccgta atgcagaaga
     4261 agaccatggg ctgggaggcc tcctccgagc ggatgtaccc cgaggacggc gccctgaagg
     4321 gcgagatcaa gcagaggctg aagctgaagg acggcggcca ctacgacgct gaggtcaaga
     4381 ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc ctacaacgtc aacatcaagt
     4441 tggacatcac ctcccacaac gaggactaca ccatcgtgga acagtacgaa cgcgccgagg
     4501 gccgccactc caccggcggc atggacgagc tgtacaagta aagcggccgc gactctagat
     4561 cataatcagc cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct
     4621 ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc
     4681 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     4741 actgcattct agttgtggtt tgtccaaact catcaatgta tctaagccgc ggtggagctc
     4801 aagtgatctc caaaaaataa gtactttttg actgtaaata aaattgtaag gagtaaaaag
     4861 tacttttttt tctaaaaaaa tgtaattaag taaaagtaaa agtattgatt tttaattgta
     4921 ctcaagtaaa gtaaaaatcc ccaaaaataa tacttaagta cagtaatcaa gtaaaattac
     4981 tcaagtactt tacaccgagc tccagctttt gttcccttta gtgagggtta attgcgcgct
     5041 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     5101 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     5161 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     5221 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     5281 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     5341 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     5401 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     5461 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     5521 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     5581 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     5641 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     5701 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     5761 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     5821 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     5881 acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     5941 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     6001 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     6061 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     6121 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     6181 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     6241 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     6301 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     6361 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     6421 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     6481 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     6541 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     6601 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6661 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6721 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6781 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     6841 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     6901 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     6961 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     7021 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     7081 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     7141 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     7201 cac
//
