LOCUS       sequence-72770-e        6940 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses human Tim3 .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6940)
  AUTHORS   Lee J, Su EW, Zhu C, Hainline S, Phuah J, Moroco JA, Smithgall TE, 
            Kuchroo VK, Kane LP
  TITLE     Phosphotyrosine-dependent coupling of Tim-3 to T-cell receptor 
            signaling pathways.
  JOURNAL   Mol Cell Biol. 2011 Oct;31(19):3963-74. doi: 10.1128/MCB.05297-11. 
            Epub 2011 Aug 1.
  PUBMED    21807895
REFERENCE   2  (bases 1 to 6940)
  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: 
            "10.1128/MCB.05297-11"; journalName: "Mol Cell Biol"; date: 
            "2011-10"; volume: "31"; issue: "19"; pages: "3963-74"
COMMENT     Sequence Label: pCDEF3‐hTIM3 (no tag)
FEATURES             Location/Qualifiers
     source          1..6940
                     /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"
     promoter        351..1529
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          582..1520
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1477..1497
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     promoter        complement(2493..2511)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2494..2511)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2531..2548)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2537..2761
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2807..3235
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2894..2913)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3104..3125
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3244..3264)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3249..3578
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3429..3564
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3491..3510
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3645..4439
                     /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(3699..3718)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4309..4328
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4613..4734
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4650..4669)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4704..4723
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4783..4799)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4783..4799)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4796..4818)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4807..4823
                     /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(4831..4837,4838..4855,4856..4861))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4876..4897
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5014..5031)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5185..5773)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5265..5284)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5944..6735,6736..6804))
                     /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     6567..6586
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6805..6909)
                     /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 taggcttttg caaaaagctt tgcaaagatg gataaagttt taaacagaga
      301 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      361 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      421 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      481 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      541 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      601 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      661 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      721 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      781 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      841 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      901 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      961 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
     1021 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
     1081 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1141 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1201 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1261 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1321 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1381 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1441 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1501 gtttttttct tccatttcag gtgtcgtgag gaattagctt ggtaccgcca ccatgttttc
     1561 acatcttccc tttgactgtg tcctgctgct gctgctgcta ctacttacaa ggtcctcaga
     1621 agtggaatac agagcggagg tcggtcagaa tgcctatctg ccctgcttct acaccccagc
     1681 cgccccaggg aacctcgtgc ccgtctgctg gggcaaagga gcctgtcctg tgtttgaatg
     1741 tggcaacgtg gtgctcagga ctgatgaaag ggatgtgaat tattggacat ccagatactg
     1801 gctaaatggg gatttccgca aaggagatgt gtccctgacc atagagaatg tgactctagc
     1861 agacagtggg atctactgct gccggatcca aatcccaggc ataatgaatg atgaaaaatt
     1921 taacctgaag ttggtcatca aaccagccaa ggtcacccct gcaccgactc tgcagagaga
     1981 cttcactgca gcctttccaa ggatgcttac caccagggga catggcccag cagagacaca
     2041 gacactgggg agcctccctg atataaatct aacacaaata tccacattgg ccaatgagtt
     2101 acgggactct agattggcca atgacttacg ggactctgga gcaaccatca gaataggcat
     2161 ctacatcgga gcagggatct gtgctgggct ggctctggct cttatcttcg gcgctttaat
     2221 tttcaaatgg tattctcata gcaaagagaa gatacagaat ttaagcctca tctctttggc
     2281 caacctccct ccctcaggat tggcaaatgc agtagcagag ggaattcgct cagaagaaaa
     2341 catctatacc attgaagaga acgtatatga agtggaggag cccaatgagt attattgcta
     2401 tgtcagcagc aggcagcaac cctcacaacc tttgggttgt cgctttgcaa tgccataggc
     2461 ggccgctcga gcatgcatct agagggccct attctatagt gtcacctaaa tgctagagct
     2521 cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     2581 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     2641 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     2701 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg
     2761 gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc
     2821 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     2881 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     2941 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     3001 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     3061 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     3121 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     3181 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     3241 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     3301 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     3361 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     3421 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     3481 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     3541 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
     3601 ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     3661 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     3721 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     3781 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc
     3841 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     3901 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     3961 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     4021 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     4081 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     4141 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg
     4201 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     4261 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     4321 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     4381 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     4441 cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt
     4501 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     4561 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
     4621 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     4681 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     4741 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     4801 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     4861 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     4921 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     4981 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     5041 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     5101 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     5161 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     5221 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     5281 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     5341 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     5401 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     5461 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     5521 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     5581 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     5641 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     5701 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     5761 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     5821 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     5881 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5941 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     6001 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     6061 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     6121 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     6181 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     6241 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     6301 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     6361 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     6421 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     6481 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     6541 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     6601 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     6661 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     6721 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     6781 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     6841 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6901 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
//
