LOCUS       sequence-251800-        7379 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  for co-expression of CTLA4 and mgSrtA on cell membrane. For 
            proof-of-concept monitoring and labeling of CD80/86 positive cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7379)
  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 7379)
  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-CTLA4-mgSrtA
FEATURES             Location/Qualifiers
     source          1..7379
                     /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             1511..1534
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2736..2753)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2742..2966
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3012..3440
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3099..3118)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3309..3330
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3449..3469)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3454..3783
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3634..3769
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3696..3715
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3850..4644
                     /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(3904..3923)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4514..4533
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4818..4939
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4855..4874)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4909..4928
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4988..5004)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4988..5004)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5001..5023)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5012..5028
                     /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(5036..5042,5043..5060,5061..5066))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5081..5102
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5219..5236)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5390..5978)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5470..5489)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6149..6940,6941..7009))
                     /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     6772..6791
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7010..7114)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7189..7208)
                     /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 gtaccgtgca cgctgctcct gctgttggcg gccgccctgg ctccgactca
     1501 gacccgcgcc gactacaagg acgacgacga caagggaagc ggactggagc aggcaaagcc
     1561 acagataccc aaggacaaga gcaaggtggc aggatacata gagataccag acgctgacat
     1621 aaaagagccc gtgtaccccg gacccgccac aagagaacaa ctgaacagag gagtgtcctt
     1681 cgccaaagaa aaccagagcc tggacgacca aaacatcagc atcgccggcc acaccttcat
     1741 cggaagaccc aactaccagt tcaccaacct gaaggccgcc aaaaagggaa gcatggtgta
     1801 cttcaaggtg ggcaacgaaa ccagaaaata caagatgacc tccataagaa acgtgaagcc
     1861 caccgccgtg ggcgtcctgg acgaacaaaa gggaaaggac aagcagctga ccctgatcac
     1921 ctgcgatgat ctgaacagag aaaccggcgt gtgggagacc agaaaaatcc tggtggccac
     1981 agaagtgaag accggtagca ccagcggcag cggcaagccc ggcagcggcg agggcagcac
     2041 caagggcagt actagtggaa gtggtaaacc tggaagtgga gatgggagta cgaaaggaac
     2101 gcgtatacag gtgacccaac cttcagtggt gttggctagc agccatggtg tcgccagctt
     2161 tccatgtgaa tattcaccat cacacaacac tgatgaggtc cgggtgactg tgctgcggca
     2221 gacaaatgac caaatgactg aggtctgtgc cacgacattc acagagaaga atacagtggg
     2281 cttcctagat taccccttct gcagtggtac ctttaatgaa agcagagtga acctcaccat
     2341 ccaaggactg agagctgttg acacgggact gtacctctgc aaggtggaac tcatgtaccc
     2401 accgccatac tttgtgggca tgggcaacgg gacgcagatt tatgtcattg atccagaacc
     2461 atgcccggat tctgacttcc tcctttggat ccttgtcgca gttagcttgg ggttgttttt
     2521 ttacagtttc ctggtcactg ctgtttcttt gagcaagatg ctaaagaaaa gaagtcctct
     2581 tacaacaggg gtctatgtga aaatgccccc aacagagcca gaatgtgaaa agcaatttca
     2641 gccttatttt attcccatca actgagaatt ctgcagatat ccagcacagt ggcggccgct
     2701 cgagtctaga gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca
     2761 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     2821 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
     2881 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
     2941 tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag
     3001 ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     3061 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     3121 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     3181 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     3241 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     3301 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     3361 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     3421 acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc
     3481 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
     3541 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     3601 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     3661 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     3721 tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     3781 aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga
     3841 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     3901 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     3961 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     4021 aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     4081 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     4141 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     4201 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     4261 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     4321 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc
     4381 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     4441 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     4501 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     4561 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     4621 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     4681 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     4741 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     4801 agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     4861 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     4921 aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt
     4981 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     5041 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     5101 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     5161 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     5221 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     5281 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     5341 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     5401 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     5461 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     5521 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     5581 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     5641 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     5701 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     5761 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     5821 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     5881 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     5941 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     6001 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     6061 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     6121 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     6181 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     6241 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     6301 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     6361 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     6421 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     6481 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     6541 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     6601 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     6661 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     6721 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     6781 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     6841 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     6901 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     6961 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     7021 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     7081 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     7141 acgtcgacgg atcgggagat ctcccgatcc cctatggtgc actctcagta caatctgctc
     7201 tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta
     7261 gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa
     7321 tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgtt
//
