LOCUS       Exported                6604 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 6604)
  TITLE     AAV plasmids
REFERENCE   2  (bases 1 to 6604)
  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..6604
                     /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..1312
                     /label=CES2A forward Gibson
                     /note="color: black; sequence: 
                     cacctgcctttctctccacaggtgtccaggcggccgcgccaccatgccattggctagac
                     ttc; 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..2988
                     /codon_start=1
                     /label=CES2A isofrom I
                     /note="color: #ff0000"
                     /translation="MPLARLPGWLCVVACGLLLLLQHVHGDYKDDDDKGGGQDSASPIR
                     NTHRGQVRGSFVHVKDTKSGVHAFLGIPFAKPPVGLLRFAPPEDPEPWSGVRDGTSQPA
                     MCLQPDIMNLEDAKEMNLILPPISMSEDCLYLNIYTPTHAQEGSNLPVMVWIHGGGLVV
                     GSASMNDVSKLAATEEIVIVAIQYRLGVLGFFSTGDQHARGNWGYLDQVAALRWVQKNI
                     AYFGGNRDRVTIFGVAAGGTSVSSHILSPMSKGLFHGAIMQSGVALLPDLISDTSEVVY
                     KTVANLSGCEATDSEALIHCLRAKSKQEILAINQVFKMIPAVVDGEFLPKHPQELLTSM
                     DFHPVPSIIGVNTDECGWGVPMFMGLDHIIKNITRETLPAVLKNTAARMMLPPECSHLL
                     VEEYMGDTEDPETLQAQFREMLGDFMFVIPALQVAHFQRSQAPVYFYEFQHLSSFIKHV
                     RPSHVKADHGDDVAFVFGSYLWDMNLDLTEEEELLKRMMMKYWANFARNGNPNSEGLPS
                     WPVLDHDEQYLQLDTQPAVGRALKARRLQFWTKTLPQKIQELKGSQDK"
     misc_feature    1294..1371
                     /label=Signal peptide
                     /note="color: #00ffff"
     misc_feature    1372..1404
                     /label=FLAG-linker
                     /note="color: #85dae9"
     misc_feature    2005..2007
                     /label=S227A
                     /note="color: #a6acb3"
     primer_bind     complement(2902..2922)
                     /label=CES2A reverse gibson
                     /note="color: black; sequence: 
                     aaaaattttgtaatccagaggttgattggatccaagctttcatctggccttcagggctc
                     gacc; added: 2021-08-05"
     CDS             2920..2925
                     /codon_start=1
                     /label=RR cleavage site
                     /note="color: #3366ff"
                     /translation="RR"
     primer_bind     complement(2961..2991)
                     /label=Primer 1
                     /note="color: black; sequence: 
                     aaactacttgtcctgagaacccttgagctcctgg; added: 2021-08-24"
     primer_bind     complement(3057..3077)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
                     /note="color: black; sequence: CATAGCGTAAAAGGAGCAACA"
     CDS             complement(3475..3486)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /note="color: #cc99b2
                     Cleavage site after base 3474"
                     /translation="IEGR"
     primer_bind     complement(3553..3570)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
                     /note="color: black; sequence: TAGAAGGCACAGTCGAGG"
     polyA_signal    3559..3766
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
                     /note="color: #a6acb3"
     repeat_region   3854..3983
                     /label=AAV2 ITR
                     /note="color: #ffcc99"
     primer_bind     complement(3900..3911)
                     /label=L4440
                     /note="L4440 vector, forward primer"
                     /note="color: black; sequence: AGCGAGTCAGTGAGCGAG"
     primer_bind     3961..3972
                     /label=L4440
                     /note="L4440 vector, forward primer"
                     /note="color: black; sequence: AGCGAGTCAGTGAGCGAG"
     primer_bind     complement(4002..4019)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
                     /note="color: black; sequence: TGTAAAACGACGGCCAGT"
     primer_bind     complement(4002..4018)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     primer_bind     complement(4011..4033)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
                     /note="color: black; sequence: CCCAGTCACGACGTTGTAAAACG"
     rep_origin      4160..4615
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     primer_bind     complement(4247..4266)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
                     /note="color: black; sequence: AGGGAAGAAAGCGAAAGGAG"
     primer_bind     4457..4478
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
                     /note="color: black; sequence: GTGGACTCTTGTTCCAAACTGG"
     promoter        4641..4745
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             4746..5606
                     /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: 4746 .. 4814 / #ccffcc / signal sequence
                        2: 4815 .. 5606 / #ccffcc
                     Cleavage site after base 4814"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4964..4983)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
                     /note="color: black; sequence: ATAATACCGCGCCACATAGC"
     rep_origin      5777..6365
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     primer_bind     6266..6285
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
                     /note="color: black; sequence: GGGAAACGCCTGGTATCTTT"
     primer_bind     6519..6536
                     /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 accatgccat tggctagact tcctggttgg
     1321 ctgtgtgttg ttgcctgtgg gctcctgctt ctcctccagc atgtgcatgg cgactacaag
     1381 gatgacgacg ataagggggg cggtcaggac tcagccagcc ccatcaggaa cacccacagg
     1441 ggtcaggtga gaggcagctt tgtccacgtg aaggacacta aatctggtgt ccatgccttc
     1501 ctgggaatcc cctttgccaa gcctcctgta ggactactgc gctttgcacc tcctgaggac
     1561 cctgagccat ggagtggtgt gagagatggg acctcacagc cggccatgtg tctgcagcct
     1621 gatataatga atttagagga tgcgaaggaa atgaatctga tcctgcctcc catctctatg
     1681 tccgaggact gcctgtatct caacatctac acaccaaccc acgcccagga gggctctaac
     1741 ctgcctgtga tggtatggat ccacggtggt ggactggttg taggatcagc ttccatgaat
     1801 gatgtatcta aactggcagc cactgaggaa atagtaattg tcgccatcca gtatcgcttg
     1861 ggggtccttg gcttcttcag cactggagac cagcatgcca gaggcaactg gggatacctg
     1921 gaccaagtgg ctgccttacg ctgggtgcag aagaatattg cttactttgg aggcaaccgt
     1981 gacagggtca ctatatttgg agtcgcagca ggaggtacaa gtgtgtcttc tcatattctg
     2041 tccccaatgt ccaaaggact ctttcatgga gccattatgc agagtggagt ggccctgctg
     2101 cctgacctca tctctgacac ttctgaggtg gtctacaaga cagtagccaa cctatctggt
     2161 tgtgaagcca cagactcaga agccctgatc cactgcctga gagcaaagag taaacaagag
     2221 atcctggcta ttaaccaggt cttcaagatg atccctgctg tggtggatgg ggagttccta
     2281 cccaagcatc ctcaggagct gctgacctct atggattttc accctgtccc cagcatcatt
     2341 ggtgtcaaca cagatgaatg tggctgggga gtccccatgt tcatgggcct tgaccatatc
     2401 ataaagaaca taaccagaga gaccctgcca gctgttctga agaacacagc agcacggatg
     2461 atgttgcctc cagaatgtag tcacctgcta gtggaggagt atatggggga cactgaggac
     2521 ccagagaccc tgcaagcaca gttcagagag atgctggggg acttcatgtt tgtgatccct
     2581 gctctccaag tggcacattt ccagcgttcc caggctcctg tctacttcta tgagttccag
     2641 catctgtcca gcttcatcaa gcatgtcagg ccatcacatg tgaaggcaga ccatggtgat
     2701 gacgttgcct ttgtctttgg ctcctatttg tgggacatga atttggacct cactgaggag
     2761 gaggagctgc taaagaggat gatgatgaag tactgggcca atttcgcaag aaatgggaac
     2821 cccaacagtg agggtctacc ctcctggcct gtgttggacc atgatgagca gtacctgcag
     2881 ctggacaccc agcctgctgt gggtcgagcc ctgaaggcca gaaggctgca gttctggacc
     2941 aagactctgc cccagaagat ccaggagctc aagggttctc aggacaagta gaagcttgga
     3001 tccaatcaac ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt
     3061 gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc
     3121 cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag
     3181 ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc
     3241 actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc
     3301 cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg
     3361 ctgttgggca ctgacaattc cgtggtgttg tcggggaaat catcgtcctt tccttggctg
     3421 ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc
     3481 ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt
     3541 cttcgagatc tgcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc
     3601 ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg
     3661 aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg
     3721 acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggactcg agttaagggc
     3781 gaattcccga taaggatctt cctagagcat ggctacgtag ataagtagca tggcgggtta
     3841 atcattaact acaaggaacc cctagtgatg gagttggcca ctccctctct gcgcgctcgc
     3901 tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc ccgggcggcc
     3961 tcagtgagcg agcgagcgcg cagccttaat taacctaatt cactggccgt cgttttacaa
     4021 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
     4081 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     4141 agcctgaatg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     4201 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     4261 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     4321 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     4381 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     4441 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     4501 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     4561 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg
     4621 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     4681 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     4741 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     4801 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     4861 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     4921 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     4981 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     5041 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     5101 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     5161 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     5221 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     5281 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     5341 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     5401 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     5461 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     5521 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     5581 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     5641 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     5701 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     5761 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     5821 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     5881 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     5941 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     6001 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     6061 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     6121 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     6181 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     6241 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     6301 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     6361 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     6421 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
     6481 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
     6541 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     6601 gctg
//
