LOCUS       sequence-259513-        5156 bp    DNA     circular SYN 15-APR-2025
DEFINITION  expresses human TSC1 (939-992) in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5156)
  AUTHORS   Qin J, Wang Z, Hoogeveen-Westerveld M, Shen G, Gong W, Nellist M, Xu
            W
  TITLE     Structural Basis of the Interaction between Tuberous Sclerosis 
            Complex 1 (TSC1) and Tre2-Bub2-Cdc16 Domain Family Member 7 
            (TBC1D7).
  JOURNAL   J Biol Chem. 2016 Apr 15;291(16):8591-601. doi: 
            10.1074/jbc.M115.701870. Epub 2016 Feb 18.
  PUBMED    26893383
REFERENCE   2  (bases 1 to 5156)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1074/jbc.M115.701870"; journalName: "J Biol Chem"; date: 
            "2016-04-15- 15"; volume: "291"; issue: "16"; pages: "8591-601"
COMMENT     Sequence Label: pCool-human TSC1 (939-992)
FEATURES             Location/Qualifiers
     source          1..5156
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    8..24
                     /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)."
     CDS             47..700
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     658..680
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             707..724
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             731..751
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     primer_bind     complement(995..1017)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1162..1180)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1248..1352
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1353..1421,1422..2213)
                     /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     complement(1571..1590)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2384..2972
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2873..2892
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3126..3143
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3216..3293
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3294..4376
                     /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(3313..3332)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(4425..4430,4431..4448,4449..4455)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4463..4479
                     /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     4487..4503
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4487..4503
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             4499..5026
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4519..4536)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4519..4535)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4528..4550)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(5128..5133,5134..5149,5150..5156)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
ORIGIN
        1 tgtggaattg tgagcggata acaatttcac acaggaaaca gtattcatgt cccctatact
       61 aggttattgg aaaattaagg gccttgtgca acccactcga cttcttttgg aatatcttga
      121 agaaaaatat gaagagcatt tgtatgagcg cgatgaaggt gataaatggc gaaacaaaaa
      181 gtttgaattg ggtttggagt ttcccaatct tccttattat attgatggtg atgttaaatt
      241 aacacagtct atggccatca tacgttatat agctgacaag cacaacatgt tgggtggttg
      301 tccaaaagag cgtgcagaga tttcaatgct tgaaggagcg gttttggata ttagatacgg
      361 tgtttcgaga attgcatata gtaaagactt tgaaactctc aaagttgatt ttcttagcaa
      421 gctacctgaa atgctgaaaa tgttcgaaga tcgtttatgt cataaaacat atttaaatgg
      481 tgatcatgta acccatcctg acttcatgtt gtatgacgct cttgatgttg ttttatacat
      541 ggacccaatg tgcctggatg cgttcccaaa attagtttgt tttaaaaaac gtattgaagc
      601 tatcccacaa attgataagt acttgaaatc cagcaagtat atagcatggc ctttgcaggg
      661 ctggcaagcc acgtttggtg gtggcgacca tcctccaaaa tcggatctgg ttccgcgtgg
      721 atctcatagc gaaaacctgt acttccaatc tcatatggga cagctgcagg ccgcagagag
      781 caggtatgag gctcagaaaa ggataaccca ggtgtttgaa ttggagatct tagatttata
      841 tggcaggttg gagaaagatg gcctcctgaa aaaacttgaa gaagaaaaag cagaagcagc
      901 tgaagcagca gaagaaaggt gagcggccgc ggatcccaat tcatcgtgac tgactgacga
      961 tctgcctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga
     1021 cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag
     1081 cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc acgtagcgat agcggagtgt
     1141 ataattcttg aagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga
     1201 taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     1261 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     1321 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     1381 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     1441 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     1501 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     1561 ttaaagttct gctatgtggc gcggtattat cccgtgttga cgccgggcaa gagcaactcg
     1621 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     1681 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     1741 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     1801 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     1861 ccataccaaa cgacgagcgt gacaccacga tgcctgcagc aatggcaaca acgttgcgca
     1921 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     1981 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     2041 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     2101 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     2161 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     2221 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     2281 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     2341 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     2401 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     2461 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     2521 caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     2581 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     2641 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     2701 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     2761 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     2821 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     2881 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     2941 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     3001 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     3061 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     3121 agcgcagcga gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta
     3181 cgcatctgtg cggtatttca caccgcataa attccgacac catcgaatgg cgcaaaacct
     3241 ttcgcggtat ggcatgatag cgcccggaag agagtcaatt cagggtggtg aatgtgaaac
     3301 cagtaacgtt atacgatgtc gcagagtatg ccggtgtctc ttatcagacc gtttcccgcg
     3361 tggtgaacca ggccagccac gtttctgcga aaacgcggga aaaagtggaa gcggcgatgg
     3421 cggagctgaa ttacattccc aaccgcgtgg cacaacaact ggcgggcaaa cagtcgttgc
     3481 tgattggcgt tgccacctcc agtctggccc tgcacgcgcc gtcgcaaatt gtcgcggcga
     3541 ttaaatctcg cgccgatcaa ctgggtgcca gcgtggtggt gtcgatggta gaacgaagcg
     3601 gcgtcgaagc ctgtaaagcg gcggtgcaca atcttctcgc gcaacgcgtc agtgggctga
     3661 tcattaacta tccgctggat gaccaggatg ccattgctgt ggaagctgcc tgcactaatg
     3721 ttccggcgtt atttcttgat gtctctgacc agacacccat caacagtatt attttctccc
     3781 atgaagacgg tacgcgactg ggcgtggagc atctggtcgc attgggtcac cagcaaatcg
     3841 cgctgttagc gggcccatta agttctgtct cggcgcgtct gcgtctggct ggctggcata
     3901 aatatctcac tcgcaatcaa attcagccga tagcggaacg ggaaggcgac tggagtgcca
     3961 tgtccggttt tcaacaaacc atgcaaatgc tgaatgaggg catcgttccc actgcgatgc
     4021 tggttgccaa cgatcagatg gcgctgggcg caatgcgcgc cattaccgag tccgggctgc
     4081 gcgttggtgc ggatatctcg gtagtgggat acgacgatac cgaagacagc tcatgttata
     4141 tcccgccgtt aaccaccatc aaacaggatt ttcgcctgct ggggcaaacc agcgtggacc
     4201 gcttgctgca actctctcag ggccaggcgg tgaagggcaa tcagctgttg cccgtctcac
     4261 tggtgaaaag aaaaaccacc ctggcgccca atacgcaaac cgcctctccc cgcgcgttgg
     4321 ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc
     4381 aacgcaatta atgtaagtta gctcactcat taggcacccc aggctttaca ctttatgctt
     4441 ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat
     4501 gaccatgatt acggattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     4561 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     4621 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggcgctt
     4681 tgcctggttt ccggcaccag aagcggtgcc ggaaagctgg ctggagtgcg atcttcctga
     4741 ggccgatact gtcgtcgtcc cctcaaactg gcagatgcac ggttacgatg cgcccatcta
     4801 caccaacgtg acctatccca ttacggtcaa tccgccgttt gttcccacgg agaatccgac
     4861 gggttgttac tcgctcacat ttaatgttga tgaaagctgg ctacaggaag gccagacgcg
     4921 aattattttt gatggcgttg gaattagctt atcgactgca cggtgcacca atgcttctgg
     4981 cgtcaggcag ccatcggaag ctgtggtatg gctgtgcagg tcgtaaatca ctgcataatt
     5041 cgtgtcgctc aaggcgcact cccgttctgg ataatgtttt ttgcgccgac atcataacgg
     5101 ttctggcaaa tattctgaaa tgagctgttg acaattaatc atcggctcgt ataatg
//
