LOCUS       sequence-215045-        7185 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of TOM1L2_VHS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7185)
  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 7185)
  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_TOM1L2_VHS
FEATURES             Location/Qualifiers
     source          1..7185
                     /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      1380..1427
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(3020..3475)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3157..3178)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3369..3388
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(3517..3539)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3625..3765
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3680..3699
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3780..3797)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3951..4538)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(4709..5500,5501..5569))
                     /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     5332..5351
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5570..5674)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5742..5760
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5783..5860
                     /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             5861..6943
                     /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(5880..5899)
                     /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 cggccatgga atttctgctg ggcaacccgt tctcaacccc ggtcggtcaa tgtctggaaa
      961 aagctacgga tggctcactg caaagcgaag actggaccct gaacatggaa atctgcgata
     1021 tcatcaacga aacggaagaa ggcccgaaag acgctatccg tgcgctgaaa aagcgcctga
     1081 acggtaatcg taactatcgc gaagtcatgc tggccctgac cgtgctggaa acgtgcgtta
     1141 agaattgtgg ccatcgtttt cacattctgg ttgcaaaccg cgatttcatc gacagtgtcc
     1201 tggtgaaaat tatcagcccg aagaacaatc cgccgaccat tgttcaggat aaagtcctgg
     1261 ccctgatcca agcgtgggcc gatgcatttc gtagctctcc ggacctgacg ggcgtggttc
     1321 atatttacga agaactgaaa cgcaagggtg tggaattccc gatggcggcc gcataataaa
     1381 ccgatacaat taaaggctcc ttttggagcc ttttttttgg agattttcaa cgtgaaaaaa
     1441 ttattattcg caattccttt agttgttcct ttctattctc actccgctga aactgttgaa
     1501 agttgtttag caaaacccca tacagaaaat tcatttacta acgtctggaa agacgacaaa
     1561 actttagatc gttacgctaa ctatgagggc tgtctgtgga atgctacagg cgttgtagtt
     1621 tgtactggtg acgaaactca gtgtctagct agagtggcgg tggctctggt tccggtgatt
     1681 ttgattatga aaagatggca aacgctaata agggggctat gaccgaaaat gccgatgaaa
     1741 acgcgctaca gtctgacgct aaaggcaaac ttgattctgt cgctactgat tacggtgctg
     1801 ctatcgatgg tttcattggt gacgtttccg gccttgctaa tggtaatggt gctactggtg
     1861 attttgctgg ctctaattcc caaatggctc aagtcggtga cggtgataat tcacctttaa
     1921 tgaataattt ccgtcaatat ttaccttccc tccctcaatc ggttgaatgt cgcccttttg
     1981 tctttggcgc tggtaaacca tatgaatttt ctattgattg tgacaaaata aacttattcc
     2041 gtggtgtctt tgcgtttctt ttatatgttg ccacctttat gtatgtattt tctacgtttg
     2101 ctaacatact gcgtaataag gagtcttaat catgccagtt cttttggcta gcgccgccct
     2161 ataccttgtc tgcctccccg cgttgcgtcg cggtgcatgg agccgggcca cctcgacctg
     2221 aatggaagcc ggcggcacct cgctaacgga ttcaccactc caagaattgg agccaatcaa
     2281 ttcttgcgga gaactgtgaa tgcgcaaacc aacccttggc agaacatatc catcgcgtcc
     2341 gccatctcca gcagccgcac gcggcgcatc tcgggcagcg ttgggtcctg gccacgggtg
     2401 cgcatgatcg tgctcctgtc gttgaggacc cggctaggct ggcggggttg ccttactggt
     2461 tagcagaatg aatcaccgat acgcgagcga acgtgaagcg actgctgctg caaaacgtct
     2521 gcgacctgag caacaacatg aatggtcttc ggtttccgtg tttcgtaaag tctggaaacg
     2581 cggaagtcag cgccctgcac cattatgttc cggatctgca tcgcaggatg ctgctggcta
     2641 ccctgtggaa cacctacatc tgtattaacg aagcgctggc attgaccctg agtgattttt
     2701 ctctggtccc gccgcatcca taccgccagt tgtttaccct cacaacgttc cagtaaccgg
     2761 gcatgttcat catcagtaac ccgtatcgtg agcatcctct ctcgtttcat cggtatcatt
     2821 acccccatga acagaaatcc cccttacacg gaggcatcag tgaccaaaca ggaaaaaacc
     2881 gcccttaaca tggcccgctt tatcagaagc cagacattaa cgcttctgga gaaactcaac
     2941 gagctggacg cggatgaaca ggcagacatc tgtgaatcgc ttcacgacca cgctgatgag
     3001 ctttaccgca ggatccgcga aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     3061 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     3121 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     3181 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     3241 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     3301 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     3361 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     3421 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcgcgg
     3481 atcctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
     3541 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
     3601 agcgggtgtt ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg atagcggagt
     3661 gtatactggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatgcgg
     3721 tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc
     3781 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     3841 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     3901 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3961 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4021 acaggactat aaagatacca gggtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     4081 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     4141 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     4201 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     4261 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     4321 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     4381 acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     4441 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     4501 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     4561 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     4621 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     4681 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     4741 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     4801 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     4861 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     4921 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     4981 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctgcaggc atcgtggtgt
     5041 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     5101 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     5161 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     5221 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     5281 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaacacgg gataataccg
     5341 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     5401 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     5461 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     5521 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     5581 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     5641 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     5701 acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc
     5761 cctttcgtct tcaagaattc ccgacaccat cgaatggtgc aaaacctttc gcggtatggc
     5821 atgatagcgc ccggaagaga gtcaattcag ggtggtgaat gtgaaaccag taacgttata
     5881 cgatgtcgca gagtatgccg gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc
     5941 cagccacgtt tctgcgaaaa cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta
     6001 cattcccaac cgcgtggcac aacaactggc gggcaaacag tcgttgctga ttggcgttgc
     6061 cacctccagt ctggccctgc acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc
     6121 cgatcaactg ggtgccagcg tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg
     6181 taaagcggcg gtgcacaatc ttctcgcgca acgcgtcagt gggctgatca ttaactatcc
     6241 gctggatgac caggatgcca ttgctgtgga agctgcctgc actaatgttc cggcgttatt
     6301 tcttgatgtc tctgaccaga cacccatcaa cagtattatt ttctcccatg aagacggtac
     6361 gcgactgggc gtggagcatc tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg
     6421 cccattaagt tctgtctcgg cgcgtctgcg tctggctggc tggcataaat atctcactcg
     6481 caatcaaatt cagccgatag cggaacggga aggcgactgg agtgccatgt ccggttttca
     6541 acaaaccatg caaatgctga atgagggcat cgttcccact gcgatgctgg ttgccaacga
     6601 tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga
     6661 tatctcggta gtgggatacg acgataccga agacagctca tgttatatcc cgccgttaac
     6721 caccatcaaa caggattttc gcctgctggg gcaaaccagc gtggaccgct tgctgcaact
     6781 ctctcagggc caggcggtga agggcaatca gctgttgccc gtctcactgg tgaaaagaaa
     6841 aaccaccctg gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
     6901 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
     6961 taagttagct cactcattag gcacaattct catgtttgac agcttatcat cgactgcacg
     7021 gtgcaccaat gcttctggcg tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc
     7081 gtaaatcact gcataattcg tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt
     7141 gcgccgacat cataacggtt ctggcaaata ttctgaaatg agctg
//
