LOCUS       Exported                6486 bp ds-DNA     circular SYN 04-MAY-2021
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6486)
  AUTHORS   Adesnik Lab
  TITLE     Direct Submission
  JOURNAL   Exported May 4, 2021 from SnapGene 4.2.11
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6486
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
                     /note="color: #ffcc99"
     protein_bind    165..183
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
                     /note="color: #31849b"
     protein_bind    202..220
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
                     /note="color: #31849b"
     protein_bind    239..257
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
                     /note="color: #31849b"
     protein_bind    276..294
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
                     /note="color: #31849b"
     protein_bind    313..331
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
                     /note="color: #31849b"
     protein_bind    350..368
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
                     /note="color: #31849b"
     protein_bind    387..405
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
                     /note="color: #31849b"
     protein_bind    424..442
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
                     /note="color: #31849b"
     primer_bind     464..488
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
                     /note="color: black; sequence: AGCTCGTTTAGTGAACCGTCAGATC"
     CDS             564..1536
                     /codon_start=1
                     /label=Chronos
                     /note="color: #a6acb3"
                     /translation="METAATMTHAFISAVPSAEATIRGLLSAAAVVTPAADAHGETSNA
                     TTAGADHGCFPHINHGTELQHKIAVGLQWFTVIVAIVQLIFYGWHSFKATTGWEEVYVC
                     VIELVKCFIELFHDVDSPATVYQTNGGAVAWLRYSEWLLTCPVILIHLSNLTGLHEEYS
                     KRTMTILVTDIGNIVWGITAAFTKGPLKILFFMIGLFYGVTCFFQIAKVYIESYHTLPK
                     GVCRKICKIMAYVFFCSWLMFPVMFIAGHEGLGLITPYTSGIGHLILDLIAKNTWGFLG
                     HHLRVKIHEHILIHGDIRKTTTINVAGENMEIETFVDEEEEGG"
     misc_feature    792..809
                     /label=seq primer 3
                     /note="color: #a6acb3"
     CDS             1537..1538
                     /codon_start=1
                     /label=Chronos
                     /note="color: #a6acb3"
                     /translation=""
     misc_feature    1539..1553
                     /label=Linker
                     /note="This feature has 2 segments:
                        1: 1539 .. 1540 / #a6acb3
                        2: 1541 .. 1553 / #a6acb3"
     CDS             1554..1577
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /note="color: #cc99b2"
                     /translation="DYKDDDDK"
     CDS             1578..1772
                     /codon_start=1
                     /label=Soma-targeting motif
                     /note="color: #339966"
                     /translation="QSQPILNTKEMAPQSKPPEELEMSSMPSPVAPLPARTEGVIDMRS
                     MSSIDSFISCATDFPEATRF"
     CDS             1788..1844
                     /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"
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1854..1874
                     /codon_start=1
                     /product="nuclear localization signal of SV40 large T 
                     antigen"
                     /label=SV40 NLS
                     /note="color: #993366"
                     /translation="PKKKRKV"
     CDS             1884..2597
                     /codon_start=1
                     /label=mRuby3
                     /note="color: #ff0000"
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTD
                     IALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAV
                     AKYSNLGGGMDELYK"
     misc_feature    2613..3201
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
                     /note="color: #ffffff"
     primer_bind     complement(2666..2686)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
                     /note="color: black; sequence: CATAGCGTAAAAGGAGCAACA"
     CDS             complement(3084..3095)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /note="color: #cc99b2
                     Cleavage site after base 3083"
                     /translation="IEGR"
     polyA_signal    3233..3709
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
                     /note="color: #a6acb3"
     primer_bind     complement(3432..3451)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
                     /note="color: black; sequence: CCAGCTTGGTTCCCAATAGA"
     repeat_region   3749..3889
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
                     /note="color: #ffe4c4"
     repeat_region   3749..3878
                     /label=AAV2 ITR
                     /note="color: #ffcc99"
     rep_origin      3964..4419
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     primer_bind     complement(4051..4070)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
                     /note="color: black; sequence: AGGGAAGAAAGCGAAAGGAG"
     primer_bind     4261..4282
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
                     /note="color: black; sequence: GTGGACTCTTGTTCCAAACTGG"
     primer_bind     complement(4436..4455)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
                     /note="color: black; sequence: CGGCATCAGAGCAGATTGTA"
     primer_bind     4555..4577
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
                     /note="color: black; sequence: CCGGGAGCTGCATGTGTCAGAGG"
     primer_bind     complement(4615..4633)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
                     /note="color: black; sequence: AATAGGCGTATCACGAGGC"
     promoter        4701..4805
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             4806..5666
                     /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: 4806 .. 4874 / #ccffcc / signal sequence
                        2: 4875 .. 5666 / #ccffcc
                     Cleavage site after base 4874"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5024..5043)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
                     /note="color: black; sequence: ATAATACCGCGCCACATAGC"
     rep_origin      5837..6425
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     primer_bind     6326..6345
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
                     /note="color: black; sequence: GGGAAACGCCTGGTATCTTT"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtatcaag tgccacctga cgtctcccta tcagtgatag
      181 agaagtcgac acgtctcgag ctccctatca gtgatagaga aggtacgtct agaacgtctc
      241 cctatcagtg atagagaagt cgacacgtct cgagctccct atcagtgata gagaaggtac
      301 gtctagaacg tctccctatc agtgatagag aagtcgacac gtctcgagct ccctatcagt
      361 gatagagaag gtacgtctag aacgtctccc tatcagtgat agagaagtcg acacgtctcg
      421 agctccctat cagtgataga gaagctaccc cctatataag cagagctcgt ttagtgaacc
      481 gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga caccgggacc
      541 gatccagcct ggaatccggt accatggaaa cagccgccac aatgacccac gcctttatct
      601 cagccgtgcc tagcgccgaa gccacaatta gaggcctgct gagcgccgca gcagtggtga
      661 caccagcagc agacgctcac ggagaaacct ctaacgccac aacagccgga gccgatcacg
      721 gttgcttccc ccacatcaac cacggaaccg agctgcagca caagatcgca gtgggactcc
      781 agtggttcac cgtgatcgtg gctatcgtgc agctcatctt ctacggttgg cacagcttca
      841 aggccacaac cggctgggag gaggtctacg tctgcgtgat cgagctcgtc aagtgcttca
      901 tcgagctgtt ccacgatgtc gacagcccag ccacagtgta ccagaccaac ggaggagccg
      961 tggcttggct gcggtacagc gagtggctcc tgacttgccc cgtgatcctg atccacctga
     1021 gcaacctgac cggactgcac gaagagtaca gcaagcggac catgaccatc ctggtgaccg
     1081 acatcggcaa catcgtgtgg gggatcacag ccgcctttac aaagggcccc ctgaagatcc
     1141 tgttcttcat gatcggcctg ttctacggcg tgacttgctt cttccagatc gccaaggtgt
     1201 atatcgagag ctaccacacc ctgcccaaag gcgtctgccg gaagatttgc aagatcatgg
     1261 cctacgtctt cttctgctct tggctgatgt tccccgtgat gttcatcgcc ggacacgagg
     1321 gactgggcct gatcacacct tacaccagcg gaatcggcca cctgatcctg gatctgatcg
     1381 ccaagaacac ttggggcttc ctgggccacc acctgagagt gaagatccac gagcacatcc
     1441 tgatccacgg cgacatccgg aagacaacca ccatcaacgt ggccggcgag aacatggaga
     1501 tcgagacctt cgtcgacgag gaggaggagg gaggaatggc ggccgcaccg gtagattaca
     1561 aagacgatga cgataagcag tcccagccca tcctcaacac caaggagatg gccccgcaga
     1621 gcaagcctcc agaggagctg gagatgagca gcatgcccag ccccgtggcc cctctgcccg
     1681 cacgcacgga gggcgtcatc gacatgcgga gcatgtccag cattgacagc ttcatcagct
     1741 gtgccacgga cttccctgaa gccaccagat tcgcggcagg aagcggagct actaacttca
     1801 gcctgctgaa gcaggctgga gacgtggagg agaaccctgg acctactagt gcacccaaga
     1861 agaagaggaa ggtggcggcc gctatggtgt ctaagggcga agagctgatc aaggaaaata
     1921 tgcgtatgaa ggtggtcatg gaaggttcgg tcaacggcca ccaattcaaa tgcacaggtg
     1981 aaggagaagg cagaccgtac gagggagtgc aaaccatgag gatcaaagtc atcgagggag
     2041 gacccctgcc atttgccttt gacattcttg ccacgtcgtt catgtatggc agccgtacct
     2101 ttatcaagta cccggccgac atccctgatt tctttaaaca gtcctttcct gagggtttta
     2161 cttgggaaag agttacgaga tacgaagatg gtggagtcgt caccgtcacg caggacacca
     2221 gccttgagga tggcgagctc gtctacaacg tcaaggtcag aggggtaaac tttccctcca
     2281 atggtcccgt gatgcagaag aagaccaagg gttgggagcc taatacagag atgatgtatc
     2341 cagcagatgg tggtctgaga ggatacactg acatcgcact gaaagttgat ggtggtggcc
     2401 atctgcactg caacttcgtg acaacttaca ggtcaaaaaa gaccgtcggg aacatcaaga
     2461 tgcccggtgt ccatgccgtt gatcaccgcc tggaaaggat cgaggagagt gacaatgaaa
     2521 cctacgtagt gcaaagagaa gtggcagttg ccaaatacag caaccttggt ggtggcatgg
     2581 acgagctgta caagtgaatt aagcttatcg ataatcaacc tctggattac aaaatttgtg
     2641 aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt
     2701 taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata
     2761 aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg
     2821 tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc
     2881 tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct
     2941 gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt
     3001 cggggaaatc atcgtccttt ccttggctgc tcgcctatgt tgccacctgg attctgcgcg
     3061 ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc
     3121 tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct
     3181 ccctttgggc cgcctccccg catcgatacc gagcgctgct cgagagatct acgggtggca
     3241 tccctgtgac ccctccccag tgcctctcct ggccctggaa gttgccactc cagtgcccac
     3301 cagccttgtc ctaataaaat taagttgcat cattttgtct gactaggtgt ccttctataa
     3361 tattatgggg tggagggggg tggtatggag caaggggcaa gttgggaaga caacctgtag
     3421 ggcctgcggg gtctattggg aaccaagctg gagtgcagtg gcacaatctt ggctcactgc
     3481 aatctccgcc tcctgggttc aagcgattct cctgcctcag cctcccgagt tgttgggatt
     3541 ccaggcatgc atgaccaggc tcagctaatt tttgtttttt tggtagagac ggggtttcac
     3601 catattggcc aggctggtct ccaactccta atctcaggtg atctacccac cttggcctcc
     3661 caaattgctg ggattacagg cgtgaaccac tgctcccttc cctgtccttc tgattttgta
     3721 ggtaaccacg tgcggaccga gcggccgcag gaacccctag tgatggagtt ggccactccc
     3781 tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc
     3841 tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg
     3901 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat
     3961 agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     4021 ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg
     4081 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     4141 ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg
     4201 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     4261 gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt
     4321 tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat
     4381 ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa
     4441 tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc
     4501 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     4561 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg
     4621 tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg
     4681 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     4741 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     4801 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     4861 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     4921 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     4981 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     5041 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     5101 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     5161 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     5221 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     5281 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     5341 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     5401 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     5461 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     5521 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     5581 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     5641 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     5701 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     5761 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     5821 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     5881 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     5941 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     6001 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     6061 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     6121 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     6181 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     6241 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     6301 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     6361 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     6421 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     6481 acatgt
//
