LOCUS       sequence-339208-        8165 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For light regulated release of transferrin receptor (TfR) from the 
            endoplasmic reticulum. Driven by CMV promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8165)
  AUTHORS   Bourke AM, Schwartz SL, Bowen AB, Kleinjan MS, Winborn CS, Kareemo 
            DJ, Gutnick A, Schwarz TL, Kennedy MJ
  TITLE     zapERtrap: A light-regulated ER release system reveals unexpected 
            neuronal trafficking pathways.
  JOURNAL   J Cell Biol. 2021 Sep 6;220(9). pii: 212461. doi: 
            10.1083/jcb.202103186. Epub 2021 Jul 9.
  PUBMED    34241635
REFERENCE   2  (bases 1 to 8165)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Cell 
            Biol."; date: "2021-09-6"; volume: "220(9). pii: 212461. doi"; 
            pages: " 10.1083/jcb.202103186. Epub 2021 Jul 9"
COMMENT     Sequence Label: TfR-mCh-DHFR
FEATURES             Location/Qualifiers
     source          1..8165
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..22
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(37..42,43..60,61..67)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    75..91
                     /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     80..102
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     99..115
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     99..115
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        144..523
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        524..727
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     678..698
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     724..748
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        958..976
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     958..975
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     989..1008
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             complement(2005..2016)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     regulatory      3361..3370
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3367..4071
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELY"
     primer_bind     complement(3512..3530)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3780..3799
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             4099..4575
                     /codon_start=1
                     /gene="folA"
                     /product="E. coli dihydrofolate reductase"
                     /label=ecDHFR
                     /translation="ISLIAALAVDRVIGMENAMPWNLPADLAWFKRNTLNKPVIMGRHT
                     WESIGRPLPGRKNIILSSQPGTDDRVTWVKSVDEAIAACGDVPEIMVIGGGRVYEQFLP
                     KAQKLYLTHIDAEVEGDTHFPDYEPDDWESVFSEFHDADAQNSHSYCFEILERR"
     regulatory      4595..4604
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    4608..4742
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4645..4664)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4699..4718
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        4811..5168
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4811..5002
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4834..4854)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5019..5154
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5188..5205)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5188..5204)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5197..5219)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5417..5872
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5504..5523)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5714..5735
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5889..5908)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6008..6030
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6068..6086)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6154..6258
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6259..6327,6328..7119)
                     /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(6477..6496)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7290..7878
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7779..7798
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8032..8049
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg
       61 tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct atgacatgat
      121 tacgaattaa ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat
      181 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
      241 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
      301 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
      361 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
      421 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
      481 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
      541 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
      601 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
      661 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
      721 cagagctcgt ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct
      781 ccatagaaga caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg
      841 gattccccgt gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt
      901 ggcttcgtta gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt
      961 taggtgacac tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag
     1021 gtccaactgc acctcggttc tatcgattga attccccggg gatccaccat ggatcaagct
     1081 agatcagcat tctctaactt gtttggtgga gaaccattgt catatacccg gttcagcctg
     1141 gctcggcaag tagatggcga taacagtcat gtggagatga aacttgctgt agatgaagaa
     1201 gaaaatgctg acaataacac aaaggccaat gtcacaaaac caaaaaggtg tagtggaagt
     1261 atctgctatg ggactattgc tgtgatcgtc tttttcttga ttggatttat gattggctac
     1321 ttgggctatt gtaaaggggt agaaccaaaa actgagtgtg agagactggc aggaaccgag
     1381 tctccagtga gggaggagcc aggagaggac ttccctgcag cacgtcgctt atattgggat
     1441 gacctgaaga gaaagttgtc ggagaaactg gacagcacag acttcaccag caccatcaag
     1501 ctgctgaatg aaaattcata tgtccctcgt gaggctggat ctcaaaaaga tgaaaatctt
     1561 gcgttgtatg ttgaaaatca atttcgtgaa tttaaactca gcaaagtctg gcgtgatcaa
     1621 cattttgtta agattcaggt caaagacagc gctcaaaact cggtgatcat agttgataag
     1681 aacggtagac ttgtttacct ggtggagaat cctgggggtt atgtggcgta tagtaaggct
     1741 gcaacagtta ctggtaaact ggtccatgct aattttggta ctaaaaaaga ttttgaggat
     1801 ttatacactc ctgtgaatgg atctatagtg attgtcagag cagggaaaat cacctttgca
     1861 gaaaaggttg caaatgctga aagcttaaat gcaattggtg tgttgatata catggaccag
     1921 actaaatttc ccattgttaa cgcagaactt tcattctttg gacatgctca tctggggaca
     1981 ggtgaccctt acacacctgg attcccttcc ttcaatcaca ctcagtttcc accatctcgg
     2041 tcatcaggat tgcctaatat acctgtccag acaatctcca gagctgctgc agaaaagctg
     2101 tttgggaata tggaaggaga ctgtccctct gactggaaaa cagactctac atgtaggatg
     2161 gtaacctcag aaagcaagaa tgtgaagctc actgtgagca atgtgctgaa agagataaaa
     2221 attcttaaca tctttggagt tattaaaggc tttgtagaac cagatcacta tgttgtagtt
     2281 ggggcccaga gagatgcatg gggccctgga gctgcaaaat ccggtgtagg cacagctctc
     2341 ctattgaaac ttgcccagat gttctcagat atggtcttaa aagatgggtt tcagcccagc
     2401 agaagcatta tctttgccag ttggagtgct ggagactttg gatcggttgg tgccactgaa
     2461 tggctagagg gatacctttc gtccctgcat ttaaaggctt tcacttatat taatctggat
     2521 aaagcggttc ttggtaccag caacttcaag gtttctgcca gcccactgtt gtatacgctt
     2581 attgagaaaa caatgcaaaa tgtgaagcat ccggttactg ggcaatttct atatcaggac
     2641 agcaactggg ccagcaaagt tgagaaactc actttagaca atgctgcttt ccctttcctt
     2701 gcatattctg gaatcccagc agtttctttc tgtttttgcg aggacacaga ttatccttat
     2761 ttgggtacca ccatggacac ctataaggaa ctgattgaga ggattcctga gttgaacaaa
     2821 gtggcacgag cagctgcaga ggtcgctggt cagttcgtga ttaaactaac ccatgatgtt
     2881 gaattgaacc tggactatga gaggtacaac agccaactgc tttcatttgt gagggatctg
     2941 aaccaataca gagcagacat aaaggaaatg ggcctgagtt tacagtggct gtattctgct
     3001 cgtggagact tcttccgtgc tacttccaga ctaacaacag atttcgggaa tgctgagaaa
     3061 acagacagat ttgtcatgaa gaaactcaat gatcgtgtca tgagagtgga gtatcacttc
     3121 ctctctccct acgtatctcc aaaagagtct cctttccgac atgtcttctg gggctccggc
     3181 tctcacacgc tgccagcttt actggagaac ttgaaactgc gtaaacaaaa taacggtgct
     3241 tttaatgaaa cgctgttcag aaaccagttg gctctagcta cttggactat tcagggagct
     3301 gcaaatgccc tctctggtga cgtttgggac attgacaatg agtttgggga tccaccggtc
     3361 gccaccatgg tgagcaaggg cgaggaggat aacatggcca tcatcaagga gttcatgcgc
     3421 ttcaaggtgc acatggaggg ctccgtgaac ggccacgagt tcgagatcga gggcgagggc
     3481 gagggccgcc cctacgaggg cacccagacc gccaagctga aggtgaccaa gggtggcccc
     3541 ctgcccttcg cctgggacat cctgtcccct cagttcatgt acggctccaa ggcctacgtg
     3601 aagcaccccg ccgacatccc cgactacttg aagctgtcct tccccgaggg cttcaagtgg
     3661 gagcgcgtga tgaacttcga ggacggcggc gtggtgaccg tgacccagga ctcctccctg
     3721 caggacggcg agttcatcta caaggtgaag ctgcgcggca ccaacttccc ctccgacggc
     3781 cccgtaatgc agaagaagac catgggctgg gaggcctcct ccgagcggat gtaccccgag
     3841 gacggcgccc tgaagggcga gatcaagcag aggctgaagc tgaaggacgg cggccactac
     3901 gacgctgagg tcaagaccac ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac
     3961 aacgtcaaca tcaagttgga catcacctcc cacaacgagg actacaccat cgtggaacag
     4021 tacgaacgcg ccgagggccg ccactccacc ggcggcatgg acgagctgta cagtgctggt
     4081 agtgctggta gtgctggtat cagtctgatt gcggcgttag cggtagatcg cgttatcggc
     4141 atggaaaacg ccatgccgtg gaacctgcct gccgatctcg cctggtttaa acgcaacacc
     4201 ttaaataaac ccgtgattat gggccgccat acctgggaat caatcggtcg tccgttgcca
     4261 ggacgcaaaa atattatcct cagcagtcaa ccgggtacgg acgatcgcgt aacgtgggtg
     4321 aagtcggtgg atgaagccat cgcggcgtgt ggtgacgtac cagaaatcat ggtgattggc
     4381 ggcggtcgcg tttatgaaca gttcttgcca aaagcgcaaa aactgtatct gacgcatatc
     4441 gacgcagaag tggaaggcga cacccatttc ccggattacg agccggatga ctgggaatcg
     4501 gtattcagcg aattccacga tgctgatgcg cagaactctc acagctattg ctttgagatt
     4561 ctggagcggc ggtaagtcga cctgcagaag cttggccgcc atggcccaac ttgtttattg
     4621 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     4681 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga
     4741 tcgggaatta attcggcgca gcaccatggc ctgaaataac ctctgaaaga ggaacttggt
     4801 taggtacctt ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa
     4861 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     4921 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     4981 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     5041 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     5101 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     5161 tttgcaaaaa gctgttaaca gcttggcact ggccgtcgtt ttacaacgtc gtgactggga
     5221 aaaccctggc gttacccaac ttaatcgcct tgcagcacat ccccctttcg ccagctggcg
     5281 taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga
     5341 atggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg
     5401 tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     5461 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     5521 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     5581 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat
     5641 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     5701 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcgggc
     5761 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     5821 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc
     5881 actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca
     5941 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     6001 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga
     6061 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct
     6121 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     6181 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     6241 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     6301 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     6361 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     6421 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     6481 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
     6541 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     6601 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     6661 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     6721 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     6781 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
     6841 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     6901 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     6961 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     7021 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     7081 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     7141 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     7201 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     7261 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     7321 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     7381 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     7441 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     7501 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     7561 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     7621 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     7681 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     7741 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     7801 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     7861 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     7921 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     7981 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
     8041 gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg
     8101 attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac
     8161 gcaat
//
