LOCUS       sequence-245283-        7532 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  for co-expression of CD40L (CD40 ligand) and mgSrtA on cell 
            membrane. For proof-of-concept monitoring and labeling of CD40 
            positive cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7532)
  AUTHORS   Ge Y, Chen L, Liu S, Zhao J, Zhang H, Chen PR
  TITLE     Enzyme-Mediated Intercellular Proximity Labeling for Detecting 
            Cell-Cell Interactions.
  JOURNAL   J Am Chem Soc. 2019 Feb 6;141(5):1833-1837. doi: 
            10.1021/jacs.8b10286. Epub 2019 Jan 29.
  PUBMED    30676735
REFERENCE   2  (bases 1 to 7532)
  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"; doi: 
            "10.1021/jacs.8b10286"; journalName: "J Am Chem Soc"; date: 
            "2019-02-6- 6"; volume: "141"; issue: "5"; pages: "1833-1837"
COMMENT     Sequence Label: pcDNA3.1-CD40L-mgSrtA
FEATURES             Location/Qualifiers
     source          1..7532
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             683..1384
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLYGMDELYK"
     primer_bind     complement(816..834)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1084..1103
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1278..1301
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     CDS             1394..1450
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2792..2815
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2889..2906)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2895..3119
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3165..3593
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3252..3271)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3462..3483
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3602..3622)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3607..3936
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3787..3922
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3849..3868
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4003..4797
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4057..4076)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4667..4686
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4971..5092
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5008..5027)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5062..5081
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5141..5157)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5141..5157)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5154..5176)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5165..5181
                     /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)."
     promoter        complement(join(5189..5195,5196..5213,5214..5219))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5234..5255
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5372..5389)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5543..6131)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5623..5642)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6302..7093,7094..7162))
                     /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     6925..6944
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7163..7267)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7342..7361)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagcgttt aaacttaagc ttatggtgag caagggcgag gaggtcatca aagagttcat
      721 gcgcttcaag gtgcgcatgg agggctccat gaacggccac gagttcgaga tcgagggcga
      781 gggcgagggc cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggcgg
      841 ccccctgccc ttcgcctggg acatcctgtc cccccagttc atgtacggct ccaaggcgta
      901 cgtgaagcac cccgccgaca tccccgatta caagaagctg tccttccccg agggcttcaa
      961 gtgggagcgc gtgatgaact tcgaggacgg cggtctggtg accgtgaccc aggactcctc
     1021 cctgcaggac ggcacgctga tctacaaggt gaagatgcgc ggcaccaact tcccccccga
     1081 cggccccgta atgcagaaga agaccatggg ctgggaggcc tccaccgagc gcctgtaccc
     1141 ccgcgacggc gtgctgaagg gcgagatcca ccaggccctg aagctgaagg acggcggcca
     1201 ctacctggtg gagttcaaga cgatctacat ggccaagaag cccgtgcaac tgcccggcta
     1261 ctactacgtg gacaccaagc tggacatcac ctcccacaac gaggactaca ccatcgtgga
     1321 acagtacgag cgctccgagg gccgccacca cctgttcctg tacggcatgg acgagctgta
     1381 caagggaagc ggagctacta acttcagcct gctgaagcag gctggagacg tggaggagaa
     1441 ccctggaccg atagaaacat acagccaacc ttcccccaga tccgtggcaa ctggacttcc
     1501 agcgagcatg aagattttta tgtatttact tactgttttc cttatcaccc aaatgattgg
     1561 atctgtgctt tttgctgtgt atcttcatag aagattggat aaggtcgaag aggaagtaaa
     1621 ccttcatgaa gattttgtat tcataaaaaa gctaaagaga tgcaacaaag gagaaggatc
     1681 tttatccttg ctgaactgtg aggagatgag aaggcaattt gaagaccttg tcaaggatat
     1741 aacgttaaac aaagaagaga aaaaagaaaa cagctttgaa atgcaaagag gtgatgagga
     1801 tcctcaaatt gcagcacacg ttgtaagcga agccaacagt aatgcagcat ccgttctaca
     1861 gtgggccaag aaaggatatt ataccatgaa aagcaacttg gtaatgcttg aaaatgggaa
     1921 acagctgacg gttaaaagag aaggactcta ttatgtctac actcaagtca ccttctgctc
     1981 taatcgggag ccttcgagtc aacgcccatt catcgtcggc ctctggctga agcccagcag
     2041 tggatctgag agaatcttac tcaaggcggc aaatacccac agttcctccc agctttgcga
     2101 gcagcagtct gttcacttgg gcggagtgtt tgaattacaa gctggtgctt ctgtgtttgt
     2161 caacgtgact gaagcaagcc aagtgatcca cagagttggc ttctcatctt ttggcttact
     2221 caaactcacc ggtagcacca gcggcagcgg caagcccggc agcggcgagg gcagcaccaa
     2281 gggcagtact agtggaagtg gtaaacctgg aagtggagat gggagtacga aaggaacgcg
     2341 tcaggcaaag ccacagatac ccaaggacaa gagcaaggtg gcaggataca tagagatacc
     2401 agacgctgac ataaaagagc ccgtgtaccc cggacccgcc acaagagaac aactgaacag
     2461 aggagtgtcc ttcgccaaag aaaaccagag cctggacgac caaaacatca gcatcgccgg
     2521 ccacaccttc atcggaagac ccaactacca gttcaccaac ctgaaggccg ccaaaaaggg
     2581 aagcatggtg tacttcaagg tgggcaacga aaccagaaaa tacaagatga cctccataag
     2641 aaacgtgaag cccaccgccg tgggcgtcct ggacgaacaa aagggaaagg acaagcagct
     2701 gaccctgatc acctgcgatg atctgaacag agaaaccggc gtgtgggaga ccagaaaaat
     2761 cctggtggcc acagaagtga agggatccgg agactacaag gatgacgatg acaaataaga
     2821 attctgcaga tatccagcac agtggcggcc gctcgagtct agagggcccg tttaaacccg
     2881 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     2941 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     3001 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     3061 caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc
     3121 ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg
     3181 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     3241 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     3301 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     3361 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     3421 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     3481 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     3541 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attaattctg
     3601 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     3661 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     3721 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     3781 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     3841 atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag
     3901 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc
     3961 attttcggat ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     4021 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     4081 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     4141 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg
     4201 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     4261 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     4321 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     4381 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     4441 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     4501 ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg
     4561 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     4621 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     4681 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     4741 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     4801 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     4861 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     4921 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     4981 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     5041 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     5101 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     5161 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     5221 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     5281 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     5341 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     5401 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     5461 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     5521 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     5581 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     5641 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     5701 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     5761 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     5821 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     5881 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     5941 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     6001 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     6061 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     6121 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     6181 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     6241 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     6301 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     6361 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     6421 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     6481 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     6541 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     6601 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     6661 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     6721 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     6781 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     6841 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     6901 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     6961 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     7021 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     7081 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     7141 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     7201 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     7261 ttccgcgcac atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga
     7321 tcccctatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct
     7381 gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac
     7441 aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc gttttgcgct
     7501 gcttcgcgat gtacgggcca gatatacgcg tt
//
