LOCUS       sequence-245262-        7146 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7146)
  AUTHORS   Levin AM, Bates DL, Ring AM, Krieg C, Lin JT, Su L, Moraga I, Raeber
            ME, Bowman GR, Novick P, Pande VS, Fathman CG, Boyman O, Garcia KC
  TITLE     Exploiting a natural conformational switch to engineer an 
            interleukin-2 'superkine'.
  JOURNAL   Nature. 2012 Mar 25;484(7395):529-33. doi: 10.1038/nature10975.
  PUBMED    22446627
REFERENCE   2  (bases 1 to 7146)
  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"; journalName: 
            "Nature"; date: "2012-03-25- 25"; volume: "484"; issue: "7395"; 
            pages: "529-33"
COMMENT     Sequence Label: IL2 pMalLP
FEATURES             Location/Qualifiers
     source          1..7146
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(131..136,137..152,153..159)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    167..183
                     /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)."
     primer_bind     172..194
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             join(253..330,331..1428)
                     /codon_start=1
                     /gene="malE"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This gene encodes a signal sequence that can direct 
                     MBP to the bacterial periplasm."
                     /translation="MKIKTGARILALSALTTMMFSASALAKIEEGKLVIWINGDKGYNG
                     LAEVGKKFEKDTGIKVTVEHPDKLEEKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAE
                     ITPDKAFQDKLYPFTWDAVRYNGKLIAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDK
                     ELKAKGKSALMFNLQEPYFTWPLIAADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLV
                     DLIKNKHMNADTDYSIAEAAFNKGETAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPS
                     KPFVGVLSAGINAASPNKELAKEFLENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKD
                     PRIAATMENAQKGEIMPNIPQMSAFWYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     1402..1425
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1477..1500
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1918..1935
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1962..1979)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1962..1978)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1971..1993)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2157..2174)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2327..2413
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2505..2532
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        2551..2642
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2643..2711,2712..3503)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2861..2880)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(3545..4058)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3740..3761)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3952..3971
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      4169..4757
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4658..4677
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4911..4928
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4943..5085
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5011..5030)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5171..5193
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5187..5378)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     promoter        5874..5951
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             5952..7034
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(5971..5990)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 aagctgtggt atggctgtgc aggtcgtaaa tcactgcata attcgtgtcg ctcaaggcgc
       61 actcccgttc tggataatgt tttttgcgcc gacatcataa cggttctggc aaatattctg
      121 aaatgagctg ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcggataa
      181 caatttcaca caggaaacag ccagtccgtt taggtgtttt cacgagcact tcaccaacaa
      241 ggaccatagc atatgaaaat aaaaacaggt gcacgcatcc tcgcattatc cgcattaacg
      301 acgatgatgt tttccgcctc ggctctcgcc aaaatcgaag aaggtaaact ggtaatctgg
      361 attaacggcg ataaaggcta taacggtctc gctgaagtcg gtaagaaatt cgagaaagat
      421 accggaatta aagtcaccgt tgagcatccg gataaactgg aagagaaatt cccacaggtt
      481 gcggcaactg gcgatggccc tgacattatc ttctgggcac acgaccgctt tggtggctac
      541 gctcaatctg gcctgttggc tgaaatcacc ccggacaaag cgttccagga caagctgtat
      601 ccgtttacct gggatgccgt acgttacaac ggcaagctga ttgcttaccc gatcgctgtt
      661 gaagcgttat cgctgattta taacaaagat ctgctgccga acccgccaaa aacctgggaa
      721 gagatcccgg cgctggataa agaactgaaa gcgaaaggta agagcgcgct gatgttcaac
      781 ctgcaagaac cgtacttcac ctggccgctg attgctgctg acgggggtta tgcgttcaag
      841 tatgaaaacg gcaagtacga cattaaagac gtgggcgtgg ataacgctgg cgcgaaagcg
      901 ggtctgacct tcctggttga cctgattaaa aacaaacaca tgaatgcaga caccgattac
      961 tccatcgcag aagctgcctt taataaaggc gaaacagcga tgaccatcaa cggcccgtgg
     1021 gcatggtcca acatcgacac cagcaaagtg aattatggtg taacggtact gccgaccttc
     1081 aagggtcaac catccaaacc gttcgttggc gtgctgagcg caggtattaa cgccgccagt
     1141 ccgaacaaag agctggcaaa agagttcctc gaaaactatc tgctgactga tgaaggtctg
     1201 gaagcggtta ataaagacaa accgctgggt gccgtagcgc tgaagtctta cgaggaagag
     1261 ttggcgaaag atccacgtat tgccgccact atggaaaacg cccagaaagg tgaaatcatg
     1321 ccgaacatcc cgcagatgtc cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc
     1381 gccagcggtc gtcagactgt cgatgaagcc ctgaaagacg cgcagactaa ttcgagctcg
     1441 aacaacaaca acaataacaa taacaacaac ctcgggctgg aagttctgtt ccaggggccc
     1501 ggagccatgg gagaattcgc acctacttca agttctacaa agaaaacaca gctacaactg
     1561 gagcatttac ttctggattt acagatgatt ttgaatggaa ttaataatta caagaatccc
     1621 aaactcacca ggatgctcac atttaagttt tacatgccca agaaggccac agaactgaaa
     1681 catcttcagt gtctagaaga agaactcaaa cctctggagg aagtgctaaa tttagctcaa
     1741 agcaaaaact ttcacttaag acccagggac ttaatcagca atatcaacgt aatagttctg
     1801 gaactaaagg gatctgaaac aacattcatg tgtgaatatg ctgatgagac agcaaccatt
     1861 gtagaatttc tgaacagatg gattaccttt tgtcaaagca tcatctcaac actaactcat
     1921 catcaccatc accattaagt cgacctgcag gcaagcttgg cactggccgt cgttttacaa
     1981 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
     2041 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     2101 agcctgaatg gcgaatggca gcttggctgt tttggcggat gagataagat tttcagcctg
     2161 atacagatta aatcagaacg cagaagcggt ctgataaaac agaatttgcc tggcggcagt
     2221 agcgcggtgg tcccacctga ccccatgccg aactcagaag tgaaacgccg tagcgccgat
     2281 ggtagtgtgg ggtctcccca tgcgagagta gggaactgcc aggcatcaaa taaaacgaaa
     2341 ggctcagtcg aaagactggg cctttcgttt tatctgttgt ttgtcggtga acgctctcct
     2401 gagtaggaca aatccgccgg gagcggattt gaacgttgcg aagcaacggc ccggagggtg
     2461 gcgggcagga cgcccgccat aaactgccag gcatcaaatt aagcagaagg ccatcctgac
     2521 ggatggcctt tttgcgtttc tacaaactct tttgtttatt tttctaaata cattcaaata
     2581 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     2641 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     2701 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     2761 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     2821 ccgaagaacg tttcccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     2881 cccgtgttga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     2941 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     3001 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     3061 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     3121 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     3181 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     3241 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     3301 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     3361 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     3421 acacgacggg gagtcaggca actatggatg aacggaatag acagatcgct gagataggtg
     3481 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     3541 atttaccccg gttgataatc agaaaagccc caaaaacagg aagattgtat aagcaaatat
     3601 ttaaattgta aacgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     3661 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     3721 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     3781 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     3841 caaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     3901 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     3961 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     4021 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtaa aaggatctag gtgaagatcc
     4081 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     4141 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     4201 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     4261 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc
     4321 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     4381 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     4441 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     4501 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     4561 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     4621 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     4681 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     4741 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     4801 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtattg
     4861 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
     4921 tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat ctgtgcggta
     4981 tttcacaccg catatatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     5041 ccagtataca ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg acacccgcca
     5101 acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct
     5161 gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg
     5221 aggcagctgc ggtaaagctc atcagcgtgg tcgtgcagcg attcacagat gtctgcctgt
     5281 tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta atgtctggct tctgataaag
     5341 cgggccatgt taagggcggt tttttcctgt ttggtcactg atgcctccgt gtaaggggga
     5401 tttctgttca tgggggtaat gataccgatg aaacgagaga ggatgctcac gatacgggtt
     5461 actgatgatg aacatgcccg gttactggaa cgttgtgagg gtaaacaact ggcggtatgg
     5521 atgcggcggg accagagaaa aatcactcag ggtcaatgcc agcgcttcgt taatacagat
     5581 gtaggtgttc cacagggtag ccagcagcat cctgcgatgc agatccggaa cataatggtg
     5641 cagggcgctg acttccgcgt ttccagactt tacgaaacac ggaaaccgaa gaccattcat
     5701 gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc ttcacgttcg ctcgcgtatc
     5761 ggtgattcat tctgctaacc agtaaggcaa ccccgccagc ctagccgggt cctcaacgac
     5821 aggagcacga tcatgcgcac ccgtggccag gacccaacgc tgcccgaaat tccgacacca
     5881 tcgaatggtg caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca
     5941 gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt
     6001 atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa
     6061 aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg
     6121 cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt
     6181 cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt
     6241 cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc
     6301 aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg
     6361 aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acacccatca
     6421 acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat ctggtcgcat
     6481 tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc
     6541 gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata gcggaacggg
     6601 aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg aatgagggca
     6661 tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca atgcgcgcca
     6721 ttaccgagtc cgggctgcgc gttggtgcgg atatctcggt agtgggatac gacgataccg
     6781 aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg
     6841 ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc
     6901 agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg
     6961 cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg
     7021 aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc tcactcatta ggcacaattc
     7081 tcatgtttga cagcttatca tcgactgcac ggtgcaccaa tgcttctggc gtcaggcagc
     7141 catcgg
//
