LOCUS       Exported                6613 bp DNA     circular SYN 28-MAR-2023
DEFINITION  AAV expression of EGFP from TBG promoter.
ACCESSION   .
VERSION     .
KEYWORDS    pAAV.TBG.PI.TurboID-KDEL.WPRE.bGH
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6613)
  TITLE     AAV plasmids
REFERENCE   2  (bases 1 to 6613)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Mar 28, 2023 from SnapGene 6.0.2
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6613
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    49..70
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
                     /note="color: #31849b"
     promoter        85..115
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This forward directional feature has 3 segments:
                        1:  85 ..  90 / #ffffff / -35
                        2:  91 .. 108 / #ffffff
                        3: 109 .. 115 / #ffffff / -10"
     protein_bind    123..139
                     /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"
     primer_bind     128..150
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
                     /note="color: black; sequence: AGCGGATAACAATTTCACACAGG"
     primer_bind     147..163
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: RIGHT"
     primer_bind     147..163
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
                     /note="color: black; sequence: CAGGAAACAGCTATGAC"
     repeat_region   201..330
                     /label=AAV2 ITR
                     /note="color: #ffcc99"
     primer_bind     complement(212..223)
                     /label=L4440
                     /note="L4440 vector, forward primer"
                     /note="color: black; sequence: AGCGAGTCAGTGAGCGAG"
     primer_bind     273..284
                     /label=L4440
                     /note="L4440 vector, forward primer"
                     /note="color: black; sequence: AGCGAGTCAGTGAGCGAG"
     promoter        670..1079
                     /label=TBG promoter
                     /note="promoter from the human thyroxine-binding globulin 
                     gene (Yan et al., 2012)"
                     /note="color: #ffffff; direction: RIGHT"
     intron          1139..1271
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
                     /note="color: #------"
     primer_bind     1181..1200
                     /label=F190
                     /note="color: black; sequence: aaactgggcttgtcgagaca; added:
                     2019-07-08"
     primer_bind     1254..1273
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
                     /note="color: black; sequence: TTGCCTTTCTCTCCACAGGT"
     primer_bind     1255..1317
                     /label=CES2C gibson forward
                     /note="color: black; sequence: 
                     cacctgcctttctctccacaggtgtccaggcggccgcgccaccATGACACGGAACCAAC
                     TACATAAC; added: 2021-08-05"
     regulatory      1283..1287
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
                     /regulatory_class="other"
     regulatory      1288..1293
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
                     /regulatory_class="other"
     CDS             1294..2997
                     /codon_start=1
                     
                     /note="color: #ff0000"
                     /translation="MTRNQLHNWLNAGFFGLLLLLIHVQGDYKDDDDKGGGQDSPEANP
                     IRNTHTGQIQGSLIHVKDTKAGVHTFLGIPFAKPPVGPLRFAPPEAPEPWSGVRDGTAH
                     PAMCLQNLDMLNEAGLPDMKMMLSSFPMSEDCLYLNIYTPAHAHEGSNLPVMVWIHGGA
                     LVIGMASMFDGSLLTVNEDLVVVTIQYRLGVLGFFSTGDQHARGNWGYLDQAAALRWVQ
                     QNIAHFGGNPDRVTIFGEAAGGTSVSSHVVSPMSQGLFHGAIMESGVALLPDLISETSE
                     MVSTTVAKLSGCEAMDSQALVRCLRGKSEAEILAINKVFKMIPAVVDGEFFPRHPKELL
                     ASEDFHPVPSIIGVNNDEFGWSIPVVMGSAQMIKGITRENLQAVLKDTAVQMMLPPECS
                     DLLMEEYMGDTEDAQTLQIQFTEMMGDFMFVIPALQVAHFQRSHAPVYFYEFQHPPSYF
                     KDVRPPHVKADHADEIPFVFASFFWGMKLDFTEEEELLSRRMMKYWANFARHGNPNSEG
                     LPYWPVMDHDEQYLQLDIQPAVGRALKAGRLQFWTKTLPQKIQELKASQDK"
     misc_feature    1294..1371
                     /label=Signal peptide
                     /note="color: #00ffff"
     misc_feature    1372..1404
                     /label=FLAG-linker
                     /note="color: #85dae9"
     misc_feature    2014..2016
                     /label=S230A
                     /note="color: #a6acb3"
     primer_bind     complement(2766..2787)
                     
                     /note="color: black; sequence: 
                     aaaaattttgtaatccagaggttgattggatccaagctttcaCCTGCTCAGCAGCTCCT
                     CCTCC; added: 2021-08-05"
     misc_feature    2785..2790
                     /label=RR cleavage site
                     /note="color: #00ffff"
     primer_bind     complement(2967..3000)
                     /label=Primer 1
                     /note="color: black; sequence: 
                     aaactaCTTGTCCTGAGAAGCCTTTAGCTCCTGGATC; added: 2021-08-24"
     primer_bind     complement(2970..3025)
                     /label=reverse
                     /note="color: black; sequence: 
                     ccagaggttgattggatccaagcttctaCTTGTCCTGAGAAGCCTTTAGCTCCTGG; 
                     added: 2021-08-25"
     primer_bind     complement(3066..3086)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
                     /note="color: black; sequence: CATAGCGTAAAAGGAGCAACA"
     CDS             complement(3484..3495)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /note="color: #cc99b2
                     Cleavage site after base 3483"
                     /translation="IEGR"
     primer_bind     complement(3562..3579)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
                     /note="color: black; sequence: TAGAAGGCACAGTCGAGG"
     polyA_signal    3568..3775
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
                     /note="color: #a6acb3"
     repeat_region   3863..3992
                     /label=AAV2 ITR
                     /note="color: #ffcc99"
     primer_bind     complement(3909..3920)
                     /label=L4440
                     /note="L4440 vector, forward primer"
                     /note="color: black; sequence: AGCGAGTCAGTGAGCGAG"
     primer_bind     3970..3981
                     /label=L4440
                     /note="L4440 vector, forward primer"
                     /note="color: black; sequence: AGCGAGTCAGTGAGCGAG"
     primer_bind     complement(4011..4028)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
                     /note="color: black; sequence: TGTAAAACGACGGCCAGT"
     primer_bind     complement(4011..4027)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     primer_bind     complement(4020..4042)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
                     /note="color: black; sequence: CCCAGTCACGACGTTGTAAAACG"
     rep_origin      4169..4624
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     primer_bind     complement(4256..4275)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
                     /note="color: black; sequence: AGGGAAGAAAGCGAAAGGAG"
     primer_bind     4466..4487
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
                     /note="color: black; sequence: GTGGACTCTTGTTCCAAACTGG"
     promoter        4650..4754
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             4755..5615
                     /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: 4755 .. 4823 / #ccffcc / signal sequence
                        2: 4824 .. 5615 / #ccffcc
                     Cleavage site after base 4823"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4973..4992)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
                     /note="color: black; sequence: ATAATACCGCGCCACATAGC"
     rep_origin      5786..6374
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     primer_bind     6275..6294
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
                     /note="color: black; sequence: GGGAAACGCCTGGTATCTTT"
     primer_bind     6528..6545
                     /label=L4440
                     /note="L4440 vector, forward primer"
                     /note="color: black; sequence: AGCGAGTCAGTGAGCGAG"
ORIGIN
        1 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta
       61 gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg
      121 aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt acgccagatt
      181 taattaaggc cttaattagg ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc
      241 ccgggcgtcg ggcgaccttt ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg
      301 gagtggccaa ctccatcact aggggttcct tgtagttaat gattaacccg ccatgctact
      361 tatctacgta gccatgctct aggaagatcg gaattcgccc ttaagctagc aggttaattt
      421 ttaaaaagca gtcaaaagtc caagtggccc ttggcagcat ttactctctc tgtttgctct
      481 ggttaataat ctcaggagca caaacattcc agatccaggt taatttttaa aaagcagtca
      541 aaagtccaag tggcccttgg cagcatttac tctctctgtt tgctctggtt aataatctca
      601 ggagcacaaa cattccagat ccggcgcgcc agggctggaa gctacctttg acatcatttc
      661 ctctgcgaat gcatgtataa tttctacaga acctattaga aaggatcacc cagcctctgc
      721 ttttgtacaa ctttccctta aaaaactgcc aattccactg ctgtttggcc caatagtgag
      781 aactttttcc tgctgcctct tggtgctttt gcctatggcc cctattctgc ctgctgaaga
      841 cactcttgcc agcatggact taaacccctc cagctctgac aatcctcttt ctcttttgtt
      901 ttacatgaag ggtctggcag ccaaagcaat cactcaaagt tcaaacctta tcattttttg
      961 ctttgttcct cttggccttg gttttgtaca tcagctttga aaataccatc ccagggttaa
     1021 tgctggggtt aatttataac taagagtgct ctagttttgc aatacaggac atgctataaa
     1081 aatggaaaga tgttgctttc tgagagactg cagaagttgg tcgtgaggca ctgggcaggt
     1141 aagtatcaag gttacaagac aggtttaagg agaccaatag aaactgggct tgtcgagaca
     1201 gagaagactc ttgcgtttct gataggcacc tattggtctt actgacatcc actttgcctt
     1261 tctctccaca ggtgtccagg cggccgcgcc accatgacac ggaaccaact acataactgg
     1321 ctgaatgctg ggttcttcgg gctgctgctt cttcttatcc atgtgcaggg tgactacaag
     1381 gatgacgacg ataagggggg cggtcaggac tcaccagagg ccaaccccat cagaaacaca
     1441 catacaggac agatccaagg cagccttatc cacgtgaagg acactaaagc tggtgtccac
     1501 accttcctgg gaatcccctt tgccaaacct cctgtaggac cactgcgatt tgcacctcct
     1561 gaggcccctg aaccatggag tggtgtgaga gatggaactg cacatccggc catgtgtctg
     1621 caaaatcttg acatgctgaa tgaagcaggc ctgccagata tgaaaatgat gctgtcttcc
     1681 ttccctatgt ctgaggactg cctgtatctc aacatctaca ccccagccca tgcccatgag
     1741 ggctctaacc tgcctgtgat ggtgtggatc catggaggtg cactggttat aggaatggct
     1801 tccatgtttg atggatctct attgacagtc aatgaggact tggtggttgt tactatccag
     1861 tatcgtctgg gtgtcctggg ctttttcagc actggagacc agcatgccag aggcaactgg
     1921 ggatacctgg accaagcagc tgccctacgc tgggtccagc agaacatcgc tcactttgga
     1981 ggcaaccctg accgggtcac tatttttggc gaggctgcag gtggcacaag tgtgtcttca
     2041 catgttgtgt cccccatgtc ccaagggctc ttccatggtg ccatcatgga gagtggggtg
     2101 gccctgcttc ctgaccttat ctctgaaacc tctgagatgg tttccactac ggtggccaag
     2161 ctttctggat gtgaggctat ggattcccag gccctggtgc gctgtctgag aggcaaaagt
     2221 gaagcagaga ttctggctat taacaaggtc ttcaagatga tccctgctgt ggtggatggg
     2281 gagttcttcc ccaggcatcc caaagagttg ttggcatctg aggattttca ccctgtcccc
     2341 agcatcattg gtgtcaacaa tgatgagttt ggctggtcca ttcctgtggt catgggctct
     2401 gctcagatga taaaaggaat aaccagagag aacctgcagg ctgttctgaa ggatacagca
     2461 gtacaaatga tgctgcctcc tgagtgtagt gacctgctaa tggaagagta catgggggac
     2521 actgaggatg cccagaccct ccaaatacag ttcacagaga tgatgggaga tttcatgttt
     2581 gtgatccctg cactccaagt agcacatttt caacgctccc atgcccctgt ctacttctat
     2641 gagttccaac atccacccag ctacttcaag gatgtcagac caccccacgt gaaggctgac
     2701 catgctgatg agattccttt tgtctttgcg tccttcttct ggggcatgaa acttgacttc
     2761 actgaggagg aggagctgct gagcaggagg atgatgaagt actgggccaa ctttgcacga
     2821 catgggaacc ccaacagtga gggtctaccc tactggcctg tgatggacca tgatgagcag
     2881 tacctgcaac tggacatcca gcctgctgtg ggtcgagccc tgaaggctgg aaggctgcag
     2941 ttctggacca agactctgcc ccagaagatc caggagctaa aggcttctca ggacaagtag
     3001 aagcttggat ccaatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt
     3061 aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct
     3121 attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt
     3181 tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac
     3241 gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct
     3301 ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca
     3361 ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt
     3421 ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc
     3481 ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct
     3541 cttccgcgtc ttcgagatct gcctcgactg tgccttctag ttgccagcca tctgttgttt
     3601 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     3661 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     3721 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggactcga
     3781 gttaagggcg aattcccgat aaggatcttc ctagagcatg gctacgtaga taagtagcat
     3841 ggcgggttaa tcattaacta caaggaaccc ctagtgatgg agttggccac tccctctctg
     3901 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     3961 cgggcggcct cagtgagcga gcgagcgcgc agccttaatt aacctaattc actggccgtc
     4021 gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca
     4081 catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa
     4141 cagttgcgca gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg
     4201 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     4261 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     4321 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     4381 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     4441 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     4501 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     4561 aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca
     4621 atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     4681 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     4741 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     4801 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     4861 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     4921 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     4981 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     5041 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     5101 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     5161 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     5221 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     5281 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     5341 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     5401 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     5461 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     5521 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     5581 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     5641 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     5701 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     5761 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     5821 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     5881 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     5941 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     6001 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     6061 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     6121 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     6181 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     6241 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     6301 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     6361 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     6421 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     6481 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     6541 gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
     6601 attaatgcag ctg
//
