LOCUS       sequence-218613-        6782 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses FKBP-CD28 in mammalian cell.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6782)
  AUTHORS   Liu Q, Zheng J, Sun W, Huo Y, Zhang L, Hao P, Wang H, Zhuang M
  TITLE     A proximity-tagging system to identify membrane protein-protein 
            interactions.
  JOURNAL   Nat Methods. 2018 Aug 13. pii: 10.1038/s41592-018-0100-5. doi: 
            10.1038/s41592-018-0100-5.
  PUBMED    30104635
REFERENCE   2  (bases 1 to 6782)
  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: "Nat 
            Methods. 2018 Aug 13. pii: 10.1038/s41592-018-0100-5. doi: 
            10.1038/s41592-018-0100-5."
COMMENT     Sequence Label: pEF6a-FKBP-CD28
FEATURES             Location/Qualifiers
     source          1..6782
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..1179
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          232..1170
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1127..1147
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     1196..1215
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1196..1214
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1323..1643
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     CDS             2314..2343
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2359..2376
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2399..2416)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2405..2629
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2675..3103
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2762..2781)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2972..2993
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3112..3132)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3117..3446
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3297..3432
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3359..3378
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3494..3541
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3560..3958
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    4116..4237
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4153..4172)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4207..4226
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4286..4302)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4286..4302)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4299..4321)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4310..4326
                     /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(4334..4340,4341..4358,4359..4364))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4379..4400
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4517..4534)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4677..5265)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4757..4776)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5436..6227,6228..6296))
                     /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     6059..6078
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6297..6401)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6476..6495)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
       61 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
      121 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
      181 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc
      241 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt
      301 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      361 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      421 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      481 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      541 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      601 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      661 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      721 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
      781 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
      841 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
      901 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
      961 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1021 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1081 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1141 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gaattagctt ggtactaata
     1201 cgactcacta tagggagacc caagctggct aggtaagctt ggtaccgagc tcggccacca
     1261 tgctcaggct gctcttggct ctcaacttat tcccttcaat tcaagtaaca ggaggcggaa
     1321 gcggagtgca ggtagaaaca atctccccgg gagatggccg cacgttcccc aagaggggac
     1381 agacctgtgt ggtgcactac accggtatgc tcgaagacgg caagaagttc gatagctccc
     1441 gagaccgaaa caagcccttc aagttcatgc tgggcaagca agaggtcata cgcggttggg
     1501 aagaaggcgt ggcccagatg agcgtagggc agcgcgccaa gctgaccatt agccccgact
     1561 acgcctacgg ggccaccggg caccccggca tcattccacc ccatgcgaca ctcgtctttg
     1621 atgtggagct gctcaagctg gaaggaggca gcggcaacaa gattttggtg aagcagtcgc
     1681 ccatgcttgt agcgtacgac aatgcggtca accttagctg caagtattcc tacaatctct
     1741 tctcaaggga gttccgggca tcccttcaca aaggactgga tagtgctgtg gaagtctgtg
     1801 ttgtatatgg gaattactcc cagcagcttc aggtttactc aaaaacgggg ttcaactgtg
     1861 atgggaaatt gggcaatgaa tcagtgacat tctacctcca gaatttgtat gttaaccaaa
     1921 cagatattta cttctgcaaa attgaagtta tgtatcctcc tccttaccta gacaatgaga
     1981 agagcaatgg aaccattatc catgtgaaag ggaaacacct ttgtccaagt cccctatttc
     2041 ccggaccttc taagcccttt tgggtgctgg tggtggttgg tggagtcctg gcttgctata
     2101 gcttgctagt aacagtggcc tttattattt tctgggtgag gagtaagagg agcaggctcc
     2161 tgcacagtga ctacatgaac atgactcccc gccgccccgg gcccacccgc aagcattacc
     2221 agccctatgc cccaccacgc gacttcgcag cctatcgctc ctagctgcag atatccagca
     2281 cagtggcggc cgctcgagtc tagagggccc ttcgaacaaa aactcatctc agaagaggat
     2341 ctgaatatgc ataccggtca tcatcaccat caccattgag tttaaacccg ctgatcagcc
     2401 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
     2461 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat
     2521 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag
     2581 gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg
     2641 gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc
     2701 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     2761 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     2821 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     2881 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     2941 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     3001 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     3061 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attaattctg tggaatgtgt
     3121 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     3181 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     3241 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     3301 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     3361 tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct
     3421 tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat
     3481 ctgatcagca cgtgttgaca attaatcatc ggcatagtat atcggcatag tataatacga
     3541 caaggtgagg aactaaacca tggccaagcc tttgtctcaa gaagaatcca ccctcattga
     3601 aagagcaacg gctacaatca acagcatccc catctctgaa gactacagcg tcgccagcgc
     3661 agctctctct agcgacggcc gcatcttcac tggtgtcaat gtatatcatt ttactggggg
     3721 accttgtgca gaactcgtgg tgctgggcac tgctgctgct gcggcagctg gcaacctgac
     3781 ttgtatcgtc gcgatcggaa atgagaacag gggcatcttg agcccctgcg gacggtgccg
     3841 acaggtgctt ctcgatctgc atcctgggat caaagccata gtgaaggaca gtgatggaca
     3901 gccgacggca gttgggattc gtgaattgct gccctctggt tatgtgtggg agggctaagc
     3961 acttcgtggc cgaggagcag gactgacacg tgctacgaga tttcgattcc accgccgcct
     4021 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc
     4081 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg
     4141 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     4201 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct
     4261 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     4321 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     4381 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     4441 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     4501 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     4561 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     4621 gaaagaacat gtgagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     4681 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     4741 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     4801 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     4861 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     4921 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     4981 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     5041 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     5101 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     5161 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     5221 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     5281 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     5341 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     5401 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     5461 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     5521 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     5581 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     5641 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     5701 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     5761 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     5821 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     5881 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     5941 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     6001 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     6061 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     6121 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     6181 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     6241 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     6301 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     6361 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     6421 ccacctgacg tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa
     6481 tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg
     6541 ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca
     6601 tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata
     6661 cgcgttgaca ttgattattg actaggcttt tgcaaaaagc tttgcaaaga tggataaagt
     6721 tttaaacaga gaggaatctt tgcagctaat ggaccttcta ggtcttgaaa ggagtgggaa
     6781 tt
//
