LOCUS       Exported                6214 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Muscle-specific expression of secretory pathway membrane-localized 
            TurboID, N-term HA tag, C-term V5 tag, in AAV expression plasmid. 
            Amp selection.
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-tMCK-mem-TurboID
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6214)
  AUTHORS   Long J
  TITLE     Jonathan Long Lab
  JOURNAL   (Unpublished)
REFERENCE   2  (bases 1 to 6214)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6214
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     intron          1004..1136
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1119..1138
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      1153..1162
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     sig_peptide     1159..1221
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
     CDS             1222..1248
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1255..2211
                     /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             2218..2259
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2263..2409
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     primer_bind     complement(2478..2498)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2896..2907)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2974..2991)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2980..3187
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3275..3404
                     /label=AAV2 ITR
     primer_bind     complement(3423..3440)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3423..3439)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3432..3454)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3581..4036
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3668..3687)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3878..3899
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4062..4166
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4167..5027
                     /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(4385..4404)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5198..5786
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5687..5706
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5940..5957
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6074..6095
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        6110..6140
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6148..6164
                     /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     6153..6175
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6172..6188
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6172..6188
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ccttaattag gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcggc
       61 ctcagtgagc gagcgagcgc gcagagaggg agtggccaac tccatcacta ggggttcctt
      121 gtagttaatg attaacccgc catgctactt atctacgtag ccatgctcta ggaagatcgg
      181 aattcgccct taagctagcc actacgggtc taggctgccc atgtaaggag gcaaggcctg
      241 gggacacccg agatgcctgg ttataattaa ccccaacacc tgctgccccc ccccccccaa
      301 cacctgctgc ctgagcctga gcggttaccc caccccggtg cctgggtctt aggctctgta
      361 caccatggag gagaagctcg ctctaaaaat aaccctgtcc ctggtgggcc cactacgggt
      421 ctaggctgcc catgtaagga ggcaaggcct ggggacaccc gagatgcctg gttataatta
      481 accccaacac ctgctgcccc ccccccccca acacctgctg cctgagcctg agcggttacc
      541 ccaccccggt gcctgggtct taggctctgt acaccatgga ggagaagctc gctctaaaaa
      601 taaccctgtc cctggtgggc cactacgggt ctaggctgcc catgtaagga ggcaaggcct
      661 ggggacaccc gagatgcctg gttataatta accccaacac ctgctgcccc ccccccccca
      721 acacctgctg cctgagcctg agcggttacc ccaccccggt gcctgggtct taggctctgt
      781 acaccatgga ggagaagctc gctctaaaaa taaccctgtc cctggtgggc ccctccctgg
      841 ggacagcccc tcctggctag tcacaccctg taggctcctc tatataaccc aggggcacag
      901 gggctgcccc cgggtcacgg ggatcctcta gacctccgaa cggtaagagc ctagcatgta
      961 gaactggtta ccctgcagaa gttggtcgtg aggcactggg caggtaagta tcaaggttac
     1021 aagacaggtt taaggagacc aatagaaact gggcttgtcg agacagagaa gactcttgcg
     1081 tttctgatag gcacctattg gtcttactga catccacttt gcctttctct ccacaggtgt
     1141 ccaggcggcc gcgccaccat ggagacagac acactcctgc tatgggtgct gctgctctgg
     1201 gttccaggtt ccactggtga ctacccatac gatgttccag attacgctgc tagcaaagac
     1261 aatactgtgc ctctgaagct gatcgctctc ctggctaatg gcgagttcca tagtggcgaa
     1321 cagctgggag aaaccctggg catgtccagg gccgctatca acaagcacat tcagactctg
     1381 cgcgactggg gcgtggacgt gttcaccgtg cccggaaagg gctactctct gcccgagcct
     1441 atcccgctgc tgaacgctaa acagattctg ggacagctgg acggcgggag cgtggcagtc
     1501 ctgcctgtgg tcgactccac caatcagtac ctgctggatc gaatcggcga gctgaagagt
     1561 ggggatgctt gcattgcaga atatcagcag gcagggagag gaagcagagg gaggaaatgg
     1621 ttctctcctt ttggagctaa cctgtacctg agtatgtttt ggcgcctgaa gcggggacca
     1681 gcagcaatcg gcctgggccc ggtcatcgga attgtcatgg cagaagcgct gcgaaagctg
     1741 ggagcagaca aggtgcgagt caaatggccc aatgacctgt atctgcagga tagaaagctg
     1801 gcaggcatcc tggtggagct ggccggaata acaggcgatg ctgcacagat cgtcattggc
     1861 gccgggatta acgtggctat gaggcgcgtg gaggaaagcg tggtcaatca gggctggatc
     1921 acactgcagg aagcagggat taacctggac aggaatactc tggccgctac gctgatccga
     1981 gagctgcggg cagccctgga actgttcgag caggaaggcc tggctccata tctgccacgg
     2041 tgggagaagc tggataactt catcaataga cccgtgaagc tgatcattgg ggacaaagag
     2101 attttcggga ttagccgggg gattgataaa cagggagccc tgctgctgga acaggacgga
     2161 gttatcaaac cctggatggg cggagaaatc agtctgcggt ctgccgaaaa gctgcagggc
     2221 aagcccatcc ccaaccccct gctgggcctg gacagcacca atgctgtggg ccaggacacg
     2281 caggaggtca tcgtggtgcc acactccttg ccctttaagg tggtggtgat ctcagccatc
     2341 ctggccctgg tggtgctcac catcatctcc cttatcatcc tcatcatgct ttggcagaag
     2401 aagccacgtt aaaagcttgg atccaatcaa cctctggatt acaaaatttg tgaaagattg
     2461 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     2521 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     2581 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     2641 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     2701 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     2761 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa
     2821 tcatcgtcct ttccttggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc
     2881 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     2941 gctctgcggc ctcttccgcg tcttcgagat ctgcctcgac tgtgccttct agttgccagc
     3001 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     3061 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     3121 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
     3181 ctggggactc gagttaaggg cgaattcccg ataaggatct tcctagagca tggctacgta
     3241 gataagtagc atggcgggtt aatcattaac tacaaggaac ccctagtgat ggagttggcc
     3301 actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc
     3361 ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagccttaa ttaacctaat
     3421 tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat
     3481 cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat
     3541 cgcccttccc aacagttgcg cagcctgaat ggcgaatggg acgcgccctg tagcggcgca
     3601 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     3661 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     3721 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     3781 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     3841 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     3901 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     3961 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     4021 ttaacgctta caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     4081 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     4141 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     4201 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     4261 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     4321 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     4381 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     4441 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     4501 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     4561 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     4621 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     4681 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     4741 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     4801 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     4861 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     4921 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     4981 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     5041 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     5101 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     5161 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     5221 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     5281 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     5341 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     5401 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     5461 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     5521 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     5581 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     5641 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     5701 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     5761 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     5821 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     5881 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     5941 gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc
     6001 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
     6061 agcgcaacgc aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta
     6121 tgcttccggc tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca
     6181 gctatgacca tgattacgcc agatttaatt aagg
//
