LOCUS       Exported                6019 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Hepatocyte-specific expression of ER-localized TurboID, N-term HA 
            tag, C-term V5 tag, in AAV expression plasmid. Amp selection.
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-TBG-ER-TurboID
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6019)
  AUTHORS   Wei W, Riley NM, Yang AC, Kim JT, Terrell SM, Li VL, 
            Garcia-Contreras M, Bertozzi CR, Long JZ.
  TITLE     Cell type-selective secretome profiling in vivo
  JOURNAL   Nature Chemical Biology. 2020
REFERENCE   2  (bases 1 to 6019)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6019
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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."
     promoter        85..115
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     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)."
     primer_bind     128..150
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     147..163
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     147..163
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   201..330
                     /label=AAV2 ITR
     promoter        670..1079
                     /label=TBG promoter
                     /note="promoter from the human thyroxine-binding globulin 
                     gene (Yan et al., 2012)"
     intron          1139..1271
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1254..1273
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      1288..1297
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1354..1380
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1387..2343
                     /codon_start=1
                     /product="mutant of the E. coli biotin ligase BioID 
                     engineered for improved catalytic efficiency (Branon et 
                     al., 2018)"
                     /label=TurboID
                     /translation="KDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRDW
                     GVDVFTVPGKGYSLPEPIPLLNAKQILGQLDGGSVAVLPVVDSTNQYLLDRIGELKSGD
                     ACIAEYQQAGRGSRGRKWFSPFGANLYLSMFWRLKRGPAAIGLGPVIGIVMAEALRKLG
                     ADKVRVKWPNDLYLQDRKLAGILVELAGITGDAAQIVIGAGINVAMRRVEESVVNQGWI
                     TLQEAGINLDRNTLAATLIRELRAALELFEQEGLAPYLPRWEKLDNFINRPVKLIIGDK
                     EIFGISRGIDKQGALLLEQDGVIKPWMGGEISLRSAEK"
     CDS             2350..2391
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(2472..2492)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2890..2901)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2968..2985)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2974..3181
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3269..3398
                     /label=AAV2 ITR
     primer_bind     complement(3417..3434)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3417..3433)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3426..3448)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3575..4030
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3662..3681)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3872..3893
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4056..4160
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4161..5021
                     /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"
     primer_bind     complement(4379..4398)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5192..5780
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5681..5700
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5934..5951
                     /label=L4440
                     /note="L4440 vector, forward primer"
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 accatggaga cagacacact cctgtgggtg
     1321 ctgctgctct gggttccagg ttccactggt gactacccat acgatgttcc agattacgct
     1381 gctagcaaag acaatactgt gcctctgaag ctgatcgctc tcctggctaa tggcgagttc
     1441 catagtggcg aacagctggg agaaaccctg ggcatgtcca gggccgctat caacaagcac
     1501 attcagactc tgcgcgactg gggcgtggac gtgttcaccg tgcccggaaa gggctactct
     1561 ctgcccgagc ctatcccgct gctgaacgct aaacagattc tgggacagct ggacggcggg
     1621 agcgtggcag tcctgcctgt ggtcgactcc accaatcagt acctgctgga tcgaatcggc
     1681 gagctgaaga gtggggatgc ttgcattgca gaatatcagc aggcagggag aggaagcaga
     1741 gggaggaaat ggttctctcc ttttggagct aacctgtacc tgagtatgtt ttggcgcctg
     1801 aagcggggac cagcagcaat cggcctgggc ccggtcatcg gaattgtcat ggcagaagcg
     1861 ctgcgaaagc tgggagcaga caaggtgcga gtcaaatggc ccaatgacct gtatctgcag
     1921 gatagaaagc tggcaggcat cctggtggag ctggccggaa taacaggcga tgctgcacag
     1981 atcgtcattg gcgccgggat taacgtggct atgaggcgcg tggaggaaag cgtggtcaat
     2041 cagggctgga tcacactgca ggaagcaggg attaacctgg acaggaatac tctggccgct
     2101 acgctgatcc gagagctgcg ggcagccctg gaactgttcg agcaggaagg cctggctcca
     2161 tatctgccac ggtgggagaa gctggataac ttcatcaata gacccgtgaa gctgatcatt
     2221 ggggacaaag agattttcgg gattagccgg gggattgata aacagggagc cctgctgctg
     2281 gaacaggacg gagttatcaa accctggatg ggcggagaaa tcagtctgcg gtctgccgaa
     2341 aagctgcagg gcaagcccat ccccaacccc ctgctgggcc tggacagcac caaagatgaa
     2401 ctctaaaagc ttggatccaa tcaacctctg gattacaaaa tttgtgaaag attgactggt
     2461 attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat
     2521 catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg
     2581 tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt
     2641 gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact
     2701 ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc
     2761 tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg
     2821 tcctttcctt ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc
     2881 tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg
     2941 cggcctcttc cgcgtcttcg agatctgcct cgactgtgcc ttctagttgc cagccatctg
     3001 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt
     3061 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg
     3121 gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg
     3181 actcgagtta agggcgaatt cccgataagg atcttcctag agcatggcta cgtagataag
     3241 tagcatggcg ggttaatcat taactacaag gaacccctag tgatggagtt ggccactccc
     3301 tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc
     3361 tttgcccggg cggcctcagt gagcgagcga gcgcgcagcc ttaattaacc taattcactg
     3421 gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt
     3481 gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct
     3541 tcccaacagt tgcgcagcct gaatggcgaa tgggacgcgc cctgtagcgg cgcattaagc
     3601 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     3661 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     3721 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     3781 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     3841 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     3901 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     3961 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     4021 cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     4081 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     4141 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     4201 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     4261 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     4321 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     4381 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     4441 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     4501 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     4561 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     4621 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     4681 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     4741 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     4801 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     4861 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     4921 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     4981 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     5041 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     5101 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     5161 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     5221 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     5281 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     5341 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     5401 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     5461 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     5521 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     5581 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     5641 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     5701 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     5761 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     5821 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     5881 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     5941 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     6001 cgattcatta atgcagctg
//
