LOCUS       sequence-464232-        6018 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Nluc-tagged PD-L1 expresion in mammalian cells; lectin-captured 
            luciferase reporter assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6018)
  AUTHORS   Chou WC, Chen WT, Kuo CT, Chang YM, Lu YS, Li CW, Hung MC, Shen CY
  TITLE     Genetic insights into carbohydrate sulfotransferase 8 and its impact
            on the immunotherapy efficacy of cancer.
  JOURNAL   Cell Rep. 2024 Jan 23;43(1):113641. doi: 
            10.1016/j.celrep.2023.113641. Epub 2023 Dec 30.
  PUBMED    38165805
REFERENCE   2  (bases 1 to 6018)
  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: "Cell Rep.";
            date: "2024-01-23"; volume: "43(1):113641. doi"; pages: " 
            10.1016/j.celrep.2023.113641. Epub 2023 Dec 30"
COMMENT     Sequence Label: pXJ-CD274-Nluc
FEATURES             Location/Qualifiers
     source          1..6018
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        2..305
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        306..509
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     460..480
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     506..530
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          671..1243
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     primer_bind     1251..1270
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             2248..2757
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     CDS             2758..2787
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     promoter        complement(2818..2836)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    3110..3231
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3147..3166)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3201..3220
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3259..3276)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3430..4018)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3510..3529)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4189..4980,4981..5049))
                     /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     4812..4831
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5050..5154)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5222..5240
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5278..5300)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5400..5419
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(5486..5941)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5623..5644)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5835..5854
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ccgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
       61 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
      121 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
      181 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
      241 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
      301 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg
      361 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
      421 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg
      481 tacggtggga ggtctatata agcagagctc gtttagtgaa ccgtcagatc gcctggagac
      541 gccatccacg ctgttttgac ctccatagaa gacaccggga ccgatccagc ctccgcggcc
      601 gggaacggtg cattggaacg gacccctagg cttttgcaaa aagctccgga tcgatcctga
      661 gaacttcagg gtgagtttgg ggacccttga ttgttctttc tttttcgcta ttgtaaaatt
      721 catgttatat ggagggggca aagttttcag ggtgttgttt agaatgggaa gatgtccctt
      781 gtatcaccat ggaccctcat gataattttg tttctttcac tttctactct gttgacaacc
      841 attgtctcct cttattttct tttcattttc tgtaactttt tcgttaaact ttagcttgca
      901 tttgtaacga atttttaaat tcacttttgt ttatttgtca gattgtaagt actttctcta
      961 atcacttttt tttcaaggca atcagggtat attatattgt acttcagcac agttttagag
     1021 aacaattgtt ataattaaat gataaggtag aatatttctg catataaatt ctggctggcg
     1081 tggaaatatt cttattggta gaaacaacta catcctggtc atcatcctgc ctttctcttt
     1141 atggttacaa tgatatacac tgtttgagat gaggataaaa tactctgagt ccaaaccggg
     1201 cccctctgct aaccatgttc atgccttctt ctttttccta cagctcctgg gcaacgtgct
     1261 ggttattgtg ctgtctcatc attttggcaa agaattgtaa tacgactcac taagggcggc
     1321 cgcggatccc ccgggctgca ggaattcgcc accatgagga tatttgctgt ctttatattc
     1381 atgacctact ggcatttgct gaacgcattt actgtcacgg ttcccaagga cctatatgtg
     1441 gtagagtatg gtagcaatat gacaattgaa tgcaaattcc cagtagaaaa acaattagac
     1501 ctggctgcac taattgtcta ttgggaaatg gaggataaga acattattca atttgtgcat
     1561 ggagaggaag acctgaaggt tcagcatagt agctacagac agagggcccg gctgttgaag
     1621 gaccagctct ccctgggaaa tgctgcactt cagatcacag atgtgaaatt gcaggatgca
     1681 ggggtgtacc gctgcatgat cagctatggt ggtgccgact acaagcgaat tactgtgaaa
     1741 gtcaatgccc catacaacaa aatcaaccaa agaattttgg ttgtggatcc agtcacctct
     1801 gaacatgaac tgacatgtca ggctgagggc taccccaagg ccgaagtcat ctggacaagc
     1861 agtgaccatc aagtcctgag tggtaagacc accaccacca attccaagag agaggagaag
     1921 cttttcaatg tgaccagcac actgagaatc aacacaacaa ctaatgagat tttctactgc
     1981 acttttagga gattagatcc tgaggaaaac catacagctg aattggtcat cccagaacta
     2041 cctctggcac atcctccaaa tgaaaggact cacttggtaa ttctgggagc catcttatta
     2101 tgccttggtg tagcactgac attcatcttc cgtttaagaa aagggagaat gatggatgtg
     2161 aaaaaatgtg gcatccaaga tacaaactca aagaagcaaa gtgatacaca tttggaggag
     2221 acgctcgagt ctggtggatc aggtggagtc ttcacactcg aagatttcgt tggggactgg
     2281 cgacagacag ccggctacaa cctggaccaa gtccttgaac agggaggtgt gtccagtttg
     2341 tttcagaatc tcggggtgtc cgtaactccg atccaaagga ttgtcctgag cggtgaaaat
     2401 gggctgaaga tcgacatcca tgtcatcatc ccgtatgaag gtctgagcgg cgaccaaatg
     2461 ggccagatcg aaaaaatttt taaggtggtg taccctgtgg atgatcatca ctttaaggtg
     2521 atcctgcact atggcacact ggtaatcgac ggggttacgc cgaacatgat cgactatttc
     2581 ggacggccgt atgaaggcat cgccgtgttc gacggcaaaa agatcactgt aacagggacc
     2641 ctgtggaacg gcaacaaaat tatcgacgag cgcctgatca accccgacgg ctccctgctg
     2701 ttccgagtaa ccatcaacgg agtgaccggc tggcggctgt gcgaacgcat tctggcggag
     2761 cagaagctga tcagcgagga ggatctgtag ggtaccaggt aagtgtaccc aattcgccct
     2821 atagtgagtc gtattacaat tcactcgatc gcccttccca acagttgcgc agcctgaatg
     2881 gcgaatggag atccaatttt taagtgtata atgtgttaaa ctactgattc taattgtttg
     2941 tgtattttag attcacagtc ccaaggctca tttcaggccc ctcagtcctc acagtctgtt
     3001 catgatcata atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc
     3061 acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat
     3121 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     3181 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt aacgcgcgac
     3241 tctagactct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     3301 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     3361 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     3421 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     3481 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     3541 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     3601 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     3661 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     3721 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     3781 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     3841 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     3901 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     3961 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     4021 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     4081 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     4141 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     4201 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     4261 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     4321 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     4381 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     4441 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     4501 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     4561 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     4621 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     4681 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     4741 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     4801 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     4861 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     4921 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     4981 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     5041 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     5101 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     5161 tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa
     5221 aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct
     5281 ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag
     5341 acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc ttaactatgc
     5401 ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg
     5461 cgtaaggaga aaataccgca tcaggaaatt gtaagcgtta atattttgtt aaaattcgcg
     5521 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     5581 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     5641 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     5701 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     5761 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     5821 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     5881 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     5941 tccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc
     6001 tattacgcca gctagagt
//
