LOCUS       sequence-214890-        8010 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of KPNA2_Arm.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8010)
  AUTHORS   Teyra J, Kelil A, Jain S, Helmy M, Jajodia R, Hooda Y, Gu J, D'Cruz 
            AA, Nicholson SE, Min J, Sudol M, Kim PM, Bader GD, Sidhu SS
  TITLE     Large-scale survey and database of high affinity ligands for peptide
            recognition modules.
  JOURNAL   Mol Syst Biol. 2020 Dec;16(12):e9310. doi: 10.15252/msb.20199310.
  PUBMED    33438817
REFERENCE   2  (bases 1 to 8010)
  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: "Mol Syst 
            Biol."; date: "2020-12"; volume: "16(12):e9310. doi"; pages: " 
            10.15252/msb.20199310"
COMMENT     Sequence Label: pHH0103_KPNA2_Arm
FEATURES             Location/Qualifiers
     source          1..8010
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..6,7..22,23..29)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    37..53
                     /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     42..64
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             138..155
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             171..824
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     782..804
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             831..854
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             867..884
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2205..2252
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(3845..4300)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3982..4003)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4194..4213
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4342..4364)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4450..4590
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4505..4524
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4605..4622)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4776..5363)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5534..6325,6326..6394))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6157..6176
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6395..6499)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6567..6585
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6608..6685
                     /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             6686..7768
                     /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(6705..6724)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca
       61 caggaaacag ccagtccgtt taggtgtttt cacgagcact tcaccaacaa ggaccataga
      121 ttatgaaaat cgaagaacac catcaccatc accattccag cggtaagctt atgtccccta
      181 tactaggtta ttggaaaatt aagggccttg tgcaacccac tcgacttctt ttggaatatc
      241 ttgaagaaaa atatgaagag catttgtatg agcgcgatga aggtgataaa tggcgaaaca
      301 aaaagtttga attgggtttg gagtttccca atcttcctta ttatattgat ggtgatgtta
      361 aattaacaca gtctatggcc atcatacgtt atatagctga caagcacaac atgttgggtg
      421 gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg agcggttttg gatattagat
      481 acggtgtttc gagaattgca tatagtaaag actttgaaac tctcaaagtt gattttctta
      541 gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt atgtcataaa acatatttaa
      601 atggtgatca tgtaacccat cctgacttca tgttgtatga cgctcttgat gttgttttat
      661 acatggaccc aatgtgcctg gatgcgttcc caaaattagt ttgttttaaa aaacgtattg
      721 aagctatccc acaaattgat aagtacttga aatccagcaa gtatatagca tggcctttgc
      781 agggctggca agccacgttt ggtggtggcg accatcctcc aaaatcggat ctggaagttc
      841 tgttccaggg gcccctgtcc agcggtctgg ttccgcgtgg ttccggtacc gcggcccagc
      901 cggccaatca gggtaccgtt aactggagtg tcgatgacat tgtgaaaggc atcaacagct
      961 ctaatgtgga aaaccagctg caagcaaccc aggcggcccg taaactgctg tcacgcgaaa
     1021 agcaaccgcc gattgataat attatccgtg cgggtctgat cccgaaattt gtttcgttcc
     1081 tgggtcgtac cgactgcagc ccgattcagt ttgaatctgc gtgggccctg acgaacatcg
     1141 ccagcggtac ctctgaacaa acgaaggcag tggttgatgg cggtgccatt ccggcattca
     1201 tctcactgct ggcttcgccg catgcgcaca tttcagaaca ggctgtgtgg gcgctgggta
     1261 atatcgcggg cgatggttcg gtttttcgtg acctggtcat taaatatggt gcagtcgatc
     1321 cgctgctggc cctgctggca gtgccggaca tgagttccct ggcctgcggc tacctgcgta
     1381 atctgacctg gacgctgagc aacctgtgtc gcaacaaaaa tccggccccg ccgattgatg
     1441 cagtggaaca gatcctgccg accctggttc gtctgctgca tcacgatgac ccggaagttc
     1501 tggctgatac ctgttgggcg atttcatatc tgacggacgg cccgaatgaa cgcatcggta
     1561 tggtcgtgaa aaccggcgtt gtcccgcagc tggttaagct gctgggtgcg tctgaactgc
     1621 cgattgtcac cccggctctg cgtgcgattg gcaacatcgt gaccggtacg gatgaacaga
     1681 cgcaagtggt tattgacgca ggcgctctgg cggtgttccc gagtctgctg accaatccga
     1741 aaacgaacat ccagaaggaa gctacctgga cgatgtccaa tattaccgcg ggtcgccagg
     1801 atcaaatcca gcaagtcgtg aaccatggcc tggttccgtt tctggttagt gtcctgtcca
     1861 aagccgattt caaaacccag aaggaagccg tgtgggcagt taccaactat acgagcggcg
     1921 gtacggttga acaaattgtc tacctggtgc actgcggtat tatcgaaccg ctgatgaatc
     1981 tgctgaccgc caaagatacg aagattatcc tggtcattct ggacgcaatt agcaacatct
     2041 tccaggcagc tgaaaaactg ggcgaaaccg aaaagctgtc tattatgatc gaagaatgtg
     2101 gcggtctgga taagatcgaa gctctgcaaa accatgaaaa cgaaagtgtg tacaaggcat
     2161 cactgtcgct gatcgaaaag tacttttccg cggccgcata ataaaccgat acaattaaag
     2221 gctccttttg gagccttttt tttggagatt ttcaacgtga aaaaattatt attcgcaatt
     2281 cctttagttg ttcctttcta ttctcactcc gctgaaactg ttgaaagttg tttagcaaaa
     2341 ccccatacag aaaattcatt tactaacgtc tggaaagacg acaaaacttt agatcgttac
     2401 gctaactatg agggctgtct gtggaatgct acaggcgttg tagtttgtac tggtgacgaa
     2461 actcagtgtc tagctagagt ggcggtggct ctggttccgg tgattttgat tatgaaaaga
     2521 tggcaaacgc taataagggg gctatgaccg aaaatgccga tgaaaacgcg ctacagtctg
     2581 acgctaaagg caaacttgat tctgtcgcta ctgattacgg tgctgctatc gatggtttca
     2641 ttggtgacgt ttccggcctt gctaatggta atggtgctac tggtgatttt gctggctcta
     2701 attcccaaat ggctcaagtc ggtgacggtg ataattcacc tttaatgaat aatttccgtc
     2761 aatatttacc ttccctccct caatcggttg aatgtcgccc ttttgtcttt ggcgctggta
     2821 aaccatatga attttctatt gattgtgaca aaataaactt attccgtggt gtctttgcgt
     2881 ttcttttata tgttgccacc tttatgtatg tattttctac gtttgctaac atactgcgta
     2941 ataaggagtc ttaatcatgc cagttctttt ggctagcgcc gccctatacc ttgtctgcct
     3001 ccccgcgttg cgtcgcggtg catggagccg ggccacctcg acctgaatgg aagccggcgg
     3061 cacctcgcta acggattcac cactccaaga attggagcca atcaattctt gcggagaact
     3121 gtgaatgcgc aaaccaaccc ttggcagaac atatccatcg cgtccgccat ctccagcagc
     3181 cgcacgcggc gcatctcggg cagcgttggg tcctggccac gggtgcgcat gatcgtgctc
     3241 ctgtcgttga ggacccggct aggctggcgg ggttgcctta ctggttagca gaatgaatca
     3301 ccgatacgcg agcgaacgtg aagcgactgc tgctgcaaaa cgtctgcgac ctgagcaaca
     3361 acatgaatgg tcttcggttt ccgtgtttcg taaagtctgg aaacgcggaa gtcagcgccc
     3421 tgcaccatta tgttccggat ctgcatcgca ggatgctgct ggctaccctg tggaacacct
     3481 acatctgtat taacgaagcg ctggcattga ccctgagtga tttttctctg gtcccgccgc
     3541 atccataccg ccagttgttt accctcacaa cgttccagta accgggcatg ttcatcatca
     3601 gtaacccgta tcgtgagcat cctctctcgt ttcatcggta tcattacccc catgaacaga
     3661 aatccccctt acacggaggc atcagtgacc aaacaggaaa aaaccgccct taacatggcc
     3721 cgctttatca gaagccagac attaacgctt ctggagaaac tcaacgagct ggacgcggat
     3781 gaacaggcag acatctgtga atcgcttcac gaccacgctg atgagcttta ccgcaggatc
     3841 cgcgaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc
     3901 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     3961 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac
     4021 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca
     4081 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg
     4141 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag
     4201 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc
     4261 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt cgcggatcct gcctcgcgcg
     4321 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     4381 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     4441 gtgtcggggc gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac
     4501 tatgcggcat cagagcagat tgtactgaga gtgcaccata tgcggtgtga aataccgcac
     4561 agatgcgtaa ggagaaaata ccgcatcagg cgctcttccg cttcctcgct cactgactcg
     4621 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     4681 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     4741 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     4801 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     4861 taccagggtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     4921 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     4981 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     5041 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     5101 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     5161 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag
     5221 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     5281 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     5341 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     5401 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     5461 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     5521 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     5581 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     5641 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     5701 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     5761 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     5821 atagtttgcg caacgttgtt gccattgctg caggcatcgt ggtgtcacgc tcgtcgtttg
     5881 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     5941 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     6001 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     6061 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     6121 ggcgaccgag ttgctcttgc ccggcgtcaa cacgggataa taccgcgcca catagcagaa
     6181 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     6241 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     6301 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     6361 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     6421 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     6481 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca
     6541 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtcttcaag
     6601 aattcccgac accatcgaat ggtgcaaaac ctttcgcggt atggcatgat agcgcccgga
     6661 agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg tcgcagagta
     6721 tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc acgtttctgc
     6781 gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc ccaaccgcgt
     6841 ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct ccagtctggc
     6901 cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc aactgggtgc
     6961 cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag cggcggtgca
     7021 caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg atgaccagga
     7081 tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg atgtctctga
     7141 ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac tgggcgtgga
     7201 gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat taagttctgt
     7261 ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc aaattcagcc
     7321 gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa ccatgcaaat
     7381 gctgaatgag ggcatcgttc ccactgcgat gctggttgcc aacgatcaga tggcgctggg
     7441 cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatatct cggtagtggg
     7501 atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca tcaaacagga
     7561 ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc agggccaggc
     7621 ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc
     7681 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     7741 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt tagctcactc
     7801 attaggcaca attctcatgt ttgacagctt atcatcgact gcacggtgca ccaatgcttc
     7861 tggcgtcagg cagccatcgg aagctgtggt atggctgtgc aggtcgtaaa tcactgcata
     7921 attcgtgtcg ctcaaggcgc actcccgttc tggataatgt tttttgcgcc gacatcataa
     7981 cggttctggc aaatattctg aaatgagctg
//
