LOCUS       sequence-240842-        7529 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 7529)
  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 7529)
  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..7529
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             917..1618
                     /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(1050..1068)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1318..1337
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1512..1535
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     CDS             1628..1684
                     /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             3026..3049
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3123..3140)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3129..3353
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3399..3827
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3486..3505)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3696..3717
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3836..3856)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3841..4170
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4021..4156
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4083..4102
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4237..5031
                     /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(4291..4310)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4901..4920
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5205..5326
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5242..5261)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5296..5315
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5375..5391)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5375..5391)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5388..5410)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5399..5415
                     /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(5423..5429,5430..5447,5448..5453))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5468..5489
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5606..5623)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5777..6362)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5857..5876)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6533..7324,7325..7393))
                     /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     7156..7175
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7394..7498)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttatgg tgagcaaggg cgaggaggtc atcaaagagt tcatgcgctt
      961 caaggtgcgc atggagggct ccatgaacgg ccacgagttc gagatcgagg gcgagggcga
     1021 gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg gcggccccct
     1081 gcccttcgcc tgggacatcc tgtcccccca gttcatgtac ggctccaagg cgtacgtgaa
     1141 gcaccccgcc gacatccccg attacaagaa gctgtccttc cccgagggct tcaagtggga
     1201 gcgcgtgatg aacttcgagg acggcggtct ggtgaccgtg acccaggact cctccctgca
     1261 ggacggcacg ctgatctaca aggtgaagat gcgcggcacc aacttccccc ccgacggccc
     1321 cgtaatgcag aagaagacca tgggctggga ggcctccacc gagcgcctgt acccccgcga
     1381 cggcgtgctg aagggcgaga tccaccaggc cctgaagctg aaggacggcg gccactacct
     1441 ggtggagttc aagacgatct acatggccaa gaagcccgtg caactgcccg gctactacta
     1501 cgtggacacc aagctggaca tcacctccca caacgaggac tacaccatcg tggaacagta
     1561 cgagcgctcc gagggccgcc accacctgtt cctgtacggc atggacgagc tgtacaaggg
     1621 aagcggagct actaacttca gcctgctgaa gcaggctgga gacgtggagg agaaccctgg
     1681 accgatagaa acatacagcc aaccttcccc cagatccgtg gcaactggac ttccagcgag
     1741 catgaagatt tttatgtatt tacttactgt tttccttatc acccaaatga ttggatctgt
     1801 gctttttgct gtgtatcttc atagaagatt ggataaggtc gaagaggaag taaaccttca
     1861 tgaagatttt gtattcataa aaaagctaaa gagatgcaac aaaggagaag gatctttatc
     1921 cttgctgaac tgtgaggaga tgagaaggca atttgaagac cttgtcaagg atataacgtt
     1981 aaacaaagaa gagaaaaaag aaaacagctt tgaaatgcaa agaggtgatg aggatcctca
     2041 aattgcagca cacgttgtaa gcgaagccaa cagtaatgca gcatccgttc tacagtgggc
     2101 caagaaagga tattatacca tgaaaagcaa cttggtaatg cttgaaaatg ggaaacagct
     2161 gacggttaaa agagaaggac tctattatgt ctacactcaa gtcaccttct gctctaatcg
     2221 ggagccttcg agtcaacgcc cattcatcgt cggcctctgg ctgaagccca gcagtggatc
     2281 tgagagaatc ttactcaagg cggcaaatac ccacagttcc tcccagcttt gcgagcagca
     2341 gtctgttcac ttgggcggag tgtttgaatt acaagctggt gcttctgtgt ttgtcaacgt
     2401 gactgaagca agccaagtga tccacagagt tggcttctca tcttttggct tactcaaact
     2461 caccggtagc accagcggca gcggcaagcc cggcagcggc gagggcagca ccaagggcag
     2521 tactagtgga agtggtaaac ctggaagtgg agatgggagt acgaaaggaa cgcgtcaggc
     2581 aaagccacag atacccaagg acaagagcaa ggtggcagga tacatagaga taccagacgc
     2641 tgacataaaa gagcccgtgt accccggacc cgccacaaga gaacaactga acagaggagt
     2701 gtccttcgcc aaagaaaacc agagcctgga cgaccaaaac atcagcatcg ccggccacac
     2761 cttcatcgga agacccaact accagttcac caacctgaag gccgccaaaa agggaagcat
     2821 ggtgtacttc aaggtgggca acgaaaccag aaaatacaag atgacctcca taagaaacgt
     2881 gaagcccacc gccgtgggcg tcctggacga acaaaaggga aaggacaagc agctgaccct
     2941 gatcacctgc gatgatctga acagagaaac cggcgtgtgg gagaccagaa aaatcctggt
     3001 ggccacagaa gtgaagggat ccggagacta caaggatgac gatgacaaat aagaattctg
     3061 cagatatcca gcacagtggc ggccgctcga gtctagaggg cccgtttaaa cccgctgatc
     3121 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     3181 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     3241 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     3301 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     3361 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     3421 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     3481 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     3541 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     3601 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     3661 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     3721 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     3781 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     3841 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3901 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     3961 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     4021 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     4081 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     4141 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     4201 ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     4261 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     4321 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     4381 gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg
     4441 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     4501 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     4561 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     4621 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     4681 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     4741 gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat
     4801 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     4861 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     4921 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     4981 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     5041 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca
     5101 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     5161 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag
     5221 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     5281 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac
     5341 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     5401 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     5461 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     5521 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     5581 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     5641 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     5701 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     5761 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     5821 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     5881 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     5941 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     6001 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     6061 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     6121 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     6181 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     6241 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     6301 ccgctggtag cggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     6361 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     6421 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     6481 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     6541 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     6601 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     6661 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     6721 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     6781 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     6841 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     6901 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     6961 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     7021 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     7081 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     7141 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     7201 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     7261 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     7321 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     7381 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     7441 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     7501 catttccccg aaaagtgcca cctgacgtc
//
