LOCUS       sequence-462858-        6566 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human HRD1 protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6566)
  AUTHORS   Zhang Z, Ren C, Xiao R, Ma S, Liu H, Dou Y, Fan Y, Wang S, Zhan P, 
            Gao C, Yue X, Li C, Gao L, Liang X, Wu Z, Ma C
  TITLE     Palmitoylation of TIM-3 promotes immune exhaustion and restrains 
            antitumor immunity.
  JOURNAL   Sci Immunol. 2024 Nov 15;9(101):eadp7302. doi: 
            10.1126/sciimmunol.adp7302. Epub 2024 Nov 15.
  PUBMED    39546589
REFERENCE   2  (bases 1 to 6566)
  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: "Sci 
            Immunol."; date: "2024-11-15"; volume: "9(101):eadp7302. doi"; 
            pages: " 10.1126/sciimmunol.adp7302. Epub 2024 Nov 15"
COMMENT     Sequence Label: Flag-HRD1
FEATURES             Location/Qualifiers
     source          1..6566
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     126..148
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     140..157
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     141..157
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        318..697
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        698..901
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     852..872
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             931..996
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    2907..3529
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3106..3125)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(3553..3573)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3558..3887
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3738..3873
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3925..3944)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3926..3944)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3958..3974)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3958..3974)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3971..3993)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3982..3998
                     /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(4006..4012,4013..4030,4031..4036))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4051..4072
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4189..4206)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4360..4948)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4440..4459)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5119..5910,5911..5979))
                     /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     5742..5761
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5980..6084)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6111..6539
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6198..6217)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6408..6429
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
       61 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
      121 gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc aagctgatct atacattgaa
      181 tcaatattgg caattagcca tattagtcat tggttatata gcataaatca atattggcta
      241 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      301 aatatgaccg ccatgttgac attgattatt gactagttat taatagtaat caattacggg
      361 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      421 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      481 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      541 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      601 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      661 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      721 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      781 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      841 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      901 tcgtttagtg aaccgtcaga attaaccatg gactacaaag accatgacgg tgattataaa
      961 gatcatgaca tcgattacaa ggatgacgat gacaagcttg cggccgcgaa ttcaatgttc
     1021 cgcacggcag tgatgatggc ggccagcctg gcgctgaccg gggctgtggt ggctcacgcc
     1081 tactacctca aacaccagtt ctaccccact gtggtgtacc tgaccaagtc cagccccagc
     1141 atggcagtcc tgtacatcca ggcctttgtc cttgtcttcc ttctgggcaa ggtgatgggc
     1201 aaggtgttct ttgggcaact gagggcagca gagatggagc accttctgga acgttcctgg
     1261 tacgccgtca cagagacttg tctggccttc accgtttttc gggatgactt cagcccccgc
     1321 tttgttgcac tcttcactct tcttctcttc ctcaaatgtt tccactggct ggctgaggac
     1381 cgtgtggact ttatggaacg cagccccaac atctcctggc tctttcactg ccgcattgtc
     1441 tctcttatgt tcctcctggg catcctggac ttcctcttcg tcagccacgc ctatcacagc
     1501 atcctgaccc gtggggcctc tgtgcagctg gtgtttggct ttgagtatgc catcctgatg
     1561 acgatggtgc tcaccatctt catcaagtat gtgctgcact ccgtggacct ccagagtgag
     1621 aacccctggg acaacaaggc tgtgtacatg ctctacacag agctgtttac aggcttcatc
     1681 aaggttctgc tgtacatggc cttcatgacc atcatgatca aggtgcacac cttcccactc
     1741 tttgccatcc ggcccatgta cctggccatg agacagttca agaaagctgt gacagatgcc
     1801 atcatgtctc gccgagccat ccgcaacatg aacaccctgt atccagatgc caccccagag
     1861 gagctccagg caatggacaa tgtctgcatc atctgccgag aagagatggt gactggtgcc
     1921 aagagactgc cctgcaacca cattttccat accagctgcc tgcgctcctg gttccagcgg
     1981 cagcagacct gccccacctg ccgtatggat gtccttcgtg catcgctgcc agcgcagtca
     2041 ccaccacccc cggagcctgc ggatcagggg ccaccccctg ccccccaccc cccaccactc
     2101 ttgcctcagc cccccaactt cccccagggc ctcctgcctc cttttcctcc aggcatgttc
     2161 ccactgtggc cccccatggg cccctttcca cctgtcccgc ctccccccag ctcaggagag
     2221 gctgtggctc ctccatccac cagtgcagca gccctttctc ggcccagtgg agcagctaca
     2281 accacagctg ctggcaccag tgctactgct gcttctgcca cagcatctgg cccaggctct
     2341 ggctctgccc cagaggctgg ccctgcccct ggtttcccct tccctcctcc ctggatgggt
     2401 atgcccctgc ctccaccctt tgccttcccc ccaatgcctg tgccccctgc gggctttgct
     2461 gggctgaccc cagaggagct acgagctctg gagggccatg agcggcagca cctggaggcc
     2521 cggctgcaga gcctgcgtaa catccacaca ctgctggacg ccgccatgct gcagatcaac
     2581 cagtacctca ccgtgctggc ctccttgggg cccccccggc ctgccacttc agtcaactcc
     2641 actgaggaga ctgccactac agttgttgct gctgcctcct ccaccagcat ccctagctca
     2701 gaggccacga ccccaacccc aggagcctcc ccaccagccc ctgaaatgga aaggcctcca
     2761 gctcctgagt cagtgggcac agaggagatg cctgaggatg gagagcccga tgcagcagag
     2821 ctccgccggc gccgcctgca gaagctggag tctcctgttg cccactgaag atctgatatc
     2881 ggtaccagtc gactctagag gatcccgggt ggcatccctg tgacccctcc ccagtgcctc
     2941 tcctggccct ggaagttgcc actccagtgc ccaccagcct tgtcctaata aaattaagtt
     3001 gcatcatttt gtctgactag gtgtccttct ataatattat ggggtggagg ggggtggtat
     3061 ggagcaaggg gcaagttggg aagacaacct gtagggcctg cggggtctat tgggaaccaa
     3121 gctggagtgc agtggcacaa tcttggctca ctgcaatctc cgcctcctgg gttcaagcga
     3181 ttctcctgcc tcagcctccc gagttgttgg gattccaggc atgcatgacc aggctcagct
     3241 aatttttgtt tttttggtag agacggggtt tcaccatatt ggccaggctg gtctccaact
     3301 cctaatctca ggtgatctac ccaccttggc ctcccaaatt gctgggatta caggcgtgaa
     3361 ccactgctcc cttccctgtc cttctgattt taaaataact ataccagcag gaggacgtcc
     3421 agacacagca taggctacct ggccatgccc aaccggtggg acatttgagt tgcttgcttg
     3481 gcactgtcct ctcatgcgtt gggtccactc agtagatgcc tgttgaattg ggtacgcggc
     3541 cagcttggct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag
     3601 gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag
     3661 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc
     3721 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     3781 atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat
     3841 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcctcgagg
     3901 aactgaaaaa ccagaaagtt aattccctat agtgagtcgt attaaattcg taatcatgtc
     3961 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     4021 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     4081 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     4141 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     4201 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     4261 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     4321 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     4381 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     4441 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     4501 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     4561 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     4621 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     4681 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     4741 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     4801 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     4861 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     4921 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     4981 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     5041 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     5101 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     5161 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     5221 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     5281 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     5341 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     5401 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     5461 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     5521 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     5581 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     5641 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     5701 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     5761 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     5821 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     5881 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     5941 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     6001 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     6061 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgcgccctg
     6121 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     6181 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     6241 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     6301 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     6361 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     6421 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     6481 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
     6541 taacaaaata ttaacgctta caattt
//
