LOCUS       Exported                7355 bp ds-DNA     circular SYN 03-AUG-2021
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7355)
  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..7355
                     /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"
     primer_bind     620..636
                     /label=Primer 1
                     /note="color: black; sequence: cgcgcgtaatacgactc; added: 
                     2021-07-22"
     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"
     primer_bind     811..840
                     /label=qPCR QUAS F
                     /note="color: black; sequence: 
                     CGAGgtcgaCAACTTTGTATAGAAAAGTTG; added: 2021-08-03"
     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..1084
                     /label=13x QUAS
                     /note="color: #a6acb3"
     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    935..950
                     /label=QUAS
                     /note="color: #a6acb3"
     misc_feature    953..968
                     /label=QUAS#
                     /note="color: #ff0000"
     misc_feature    973..988
                     /label=QUAS
                     /note="color: #a6acb3"
     misc_feature    993..1008
                     /label=QUAS
                     /note="color: #a6acb3"
     misc_feature    1011..1026
                     /label=QUAS#
                     /note="color: #ff0000"
     misc_feature    1031..1046
                     /label=QUAS
                     /note="color: #a6acb3"
     misc_feature    1051..1066
                     /label=QUAS
                     /note="color: #a6acb3"
     misc_feature    1069..1084
                     /label=QUAS
                     /note="color: #a6acb3"
     misc_feature    1126..1158
                     /label=minimal promoter
                     /note="color: #a6acb3"
     primer_bind     complement(1170..1193)
                     /label=T4 Mlu1 10x QUAS-zsG R
                     /note="color: black; sequence: TACAaACTTGACGCGTCTTCGAGG; 
                     added: 2021-08-03"
     protein_bind    1183..1207
                     /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             1214..1237
                     /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: 1214 .. 1222 / #cc99b2
                        2: 1223 .. 1237 / #cc99b2 / enterokinase site
                     Cleavage site after base 1237"
                     /translation="DLYDDDDK"
     CDS             1238..1927
                     /codon_start=1
                     /product="Zoanthus green fluorescent protein"
                     /label=ZsGreen1
                     /note="mammalian codon-optimized"
                     /note="color: #05fd14"
                     /translation="AQSKHGLTKEMTMKYRMEGCVDGHKFVITGEGIGYPFKGKQAINL
                     CVVEGGPLPFAEDILSAAFMYGNRVFTEYPQDIVDYFKNSCPAGYTWDRSFLFEDGAVC
                     ICNADITVSVEENCMYHESKFYGVNFPADGPVMKKMTDNWEPSCEKIIPVPKQGILKGD
                     VSMYLLLKDGGRLRCQFDTVYKAKSVPRKMPDWHFIQHKLTREDRSDAKNQKWHLTEHA
                     IASGSALP"
     misc_feature    1294..1306
                     /label=Feature 40
                     /note="color: #a6acb3"
     CDS             1943..1999
                     /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 1996"
                     /translation="ATNFSLLKQAGDVEENPGP"
     primer_bind     2006..2022
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: RIGHT"
     misc_feature    2062..3266
                     /label=Ocean pout terminator
                     /note="color: #ff0000"
     misc_feature    3275..3982
                     /label=Crystallin alpha promoter
                     /note="color: #ff00ff"
     regulatory      3977..3986
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
     CDS             3983..4693
                     /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"
     polyA_signal    4816..4897
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     misc_feature    4947..5154
                     /label=minitol2-R
                     /note="color: #ff0000"
     promoter        complement(5167..5185)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     primer_bind     complement(5206..5222)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     protein_bind    5230..5246
                     /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(5254..5284)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This reverse directional feature has 3 segments:
                        1: 5254 .. 5260 / #ffffff / -10
                        2: 5261 .. 5278 / #ffffff
                        3: 5279 .. 5284 / #ffffff / -35"
     protein_bind    5299..5320
                     /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(5608..6196)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     CDS             complement(6367..7227)
                     /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: 6367 .. 7158 / #ccffcc
                        2: 7159 .. 7227 / #ccffcc / signal sequence
                     Cleavage site after base 7158"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(7228..7332)
                     /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 cctcgggtaa tcgcttatcc tcggataaac
      961 aattatcctc acgggtaatc gcttatccgc tcgggtaatc gcttatcctc ggataaacaa
     1021 ttatcctcac gggtaatcgc ttatccgctc gggtaatcgc ttatcctcgg gtaatcgctt
     1081 atccttgcaa gggtcgactc tagagggtat ataatggatc ccatcgcgtc tcagcctcac
     1141 tttgagctcc tccacacgaa ttccctcgac ctcgaagacg cgtcaagttt gtacaaaaaa
     1201 gcaggtcacc atggatctgt acgacgatga cgataaggcc cagtccaagc acggcctgac
     1261 caaggagatg accatgaagt accgcatgga gggctgcgtg gacggccaca agttcgtgat
     1321 caccggcgag ggcatcggct accccttcaa gggcaagcag gccatcaacc tgtgcgtggt
     1381 ggagggcggc cccttgccct tcgccgagga catcttgtcc gccgccttca tgtacggcaa
     1441 ccgcgtgttc accgagtacc cccaggacat cgtcgactac ttcaagaact cctgccccgc
     1501 cggctacacc tgggaccgct ccttcctgtt cgaggacggc gccgtgtgca tctgcaacgc
     1561 cgacatcacc gtgagcgtgg aggagaactg catgtaccac gagtccaagt tctacggcgt
     1621 gaacttcccc gccgacggcc ccgtgatgaa gaagatgacc gacaactggg agccctcctg
     1681 cgagaagatc atccccgtgc ccaagcaggg catcttgaag ggcgacgtga gcatgtacct
     1741 gctgctgaag gacggtggcc gcttgcgctg ccagttcgac accgtgtaca aggccaagtc
     1801 cgtgccccgc aagatgcccg actggcactt catccagcac aagctgaccc gcgaggaccg
     1861 cagcgacgcc aagaaccaga agtggcacct gaccgagcac gccatcgcct ccggctccgc
     1921 cttgcccgca tgcggaagcg gagctactaa cttcagcctg ctgaagcagg ctggagacgt
     1981 ggaggagaac cctggaccta gatctcgagg tcgacggtat cgataagctt gatatcgaat
     2041 tcctgcagcc cgggggatcc actagtaatt aagtctcagc caccgttaac tgaacatgtc
     2101 aaaacctgtg gagactgttg agatttgatg ttctgaaaag ataaagccta taaataaaat
     2161 gttgcccaaa tttcctgcct gatgtttttc tttgtctttg ctacatggct ttgctgctcg
     2221 gatcggctca ctctgtgtat gccacgttca ctttgtactc tccttctcac ggtaggttta
     2281 ttatttttag atgtgcagtt agtttctgtg aaataacaca ccacacactg atattgtctg
     2341 tgcattgact tggtgagtgc acattgtttt tgatcttgac atatttatat ttgattgatc
     2401 aggtgaactg tgtgaatcta aagtgctcca tacagatgtt ctgcattgaa aatattctca
     2461 ttttattagt ggaagtgagt gtatgccaca tccaatcaat ttcagcaaac accccagtat
     2521 gatttaatgc aaaaaaatga aggtatcaaa cacgcattac tactttgcag ttaaatattt
     2581 aacatttatt ccaacacgaa aaaaagcagt aaataacact ttgacaaaca cgtcaggaca
     2641 tcttattttt gtcaccctca caggcaattt agtataatat attatatata tatatatatc
     2701 atataataat attcagtata atatatatat atatatcata ttataatatt cagtataata
     2761 taaaacacaa acacatatat gtataatata atataacatt tttatttatt gagatgcctc
     2821 tatggaccgt gttataagaa gtaaagatca ggagaagtaa acatgaagtg taattatgaa
     2881 tactgatgtt aaattaagct atgatgagtt ttcactgtta atttaccatc tcaattaaat
     2941 gttgatgcct ccatgaccaa gttaagcaga tgagactgag acaactgtag aagacaagat
     3001 gttcactttg ctgaatatag ctggcttgac agttatctat gactctataa atatatatat
     3061 attttttttt ttataaaatg atttatttat aactatatat ccatttctca gacaggtgct
     3121 tcatatccct cactcccgta gctgtccatg ctggatctgt ccccgttgtt tttaaaaagc
     3181 taaataagtt attaacatga ctgcatccag cgagccaaac ctgtctggtg tacagctacc
     3241 agagaagctt gagatcctag tcaccggcgg ccgctaatat ggcaataaac gatcttcaga
     3301 gaaacttgca ttggtaaata gtttcagcct atgcatatgt agttattcaa actgtaatga
     3361 ctatcttaca gtattatatg tttataatag tttgatccaa taaatccaga tttatatgct
     3421 tttcaacata aaattgtgaa atgcacagaa tgtacatata caatacattt cctacaataa
     3481 tgctctcttt tctattaata gtgtgcattc agtgcaggat gttatgtatt tattttctgt
     3541 agtgcaaact ttttcaaaga attttccagt ctaagctttt cagtcaagaa aaaaaaaagc
     3601 gattgattat ctgtgatatt taaaaaaatt agctcttaaa tctggtgaac caagtggcca
     3661 caacacatta aatactgacg ttctacagcc atgtagcttc atctggtctg gtttgttttg
     3721 gcaggcactt attttccggt gatagttggc cacaagagta ttgtctgagc cattcagtgc
     3781 tagactgtca ttctcaggtc agagtccacc ccgctgattc tgctgacaac actgttccca
     3841 ccatgagata atgccattcc agagagatcc atttgtaagc ccctctttct gcagcacagg
     3901 tatataacca ggggtctgcc tccactaagg ccggcacaca tcatttgggg atctttgtac
     3961 tgtggatcca ccggtcgcca ccatggtgag caagggcgag gaggataaca tggccatcat
     4021 caaggagttc atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga
     4081 gatcgagggc gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt
     4141 gaccaagggt ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg
     4201 ctccaaggcc tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc
     4261 cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac
     4321 ccaggactcc tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa
     4381 cttcccctcc gacggccccg taatgcagaa gaagaccatg ggctgggagg cctcctccga
     4441 gcggatgtac cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa
     4501 ggacggcggc cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca
     4561 gctgcccggc gcctacaacg tcaacatcaa gttggacatc acctcccaca acgaggacta
     4621 caccatcgtg gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga
     4681 gctgtacaag taaagcggcc gcgactctag atcataatca gccataccac atttgtagag
     4741 gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat
     4801 gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     4861 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     4921 ctcatcaatg tatctaagcc gcggtggagc tcaagtgatc tccaaaaaat aagtactttt
     4981 tgactgtaaa taaaattgta aggagtaaaa agtacttttt tttctaaaaa aatgtaatta
     5041 agtaaaagta aaagtattga tttttaattg tactcaagta aagtaaaaat ccccaaaaat
     5101 aatacttaag tacagtaatc aagtaaaatt actcaagtac tttacaccga gctccagctt
     5161 ttgttccctt tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc
     5221 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     5281 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     5341 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     5401 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     5461 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     5521 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     5581 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     5641 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     5701 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     5761 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     5821 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     5881 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     5941 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     6001 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg
     6061 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     6121 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     6181 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     6241 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     6301 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     6361 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     6421 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     6481 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     6541 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     6601 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     6661 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     6721 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     6781 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     6841 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     6901 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6961 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     7021 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     7081 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     7141 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     7201 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     7261 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     7321 aggggttccg cgcacatttc cccgaaaagt gccac
//
