LOCUS       sequence-468759-        5601 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression of mouse RIT1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5601)
  AUTHORS   Fritsch R, de Krijger I, Fritsch K, George R, Reason B, Kumar MS, 
            Diefenbacher M, Stamp G, Downward J
  TITLE     RAS and RHO families of GTPases directly regulate distinct 
            phosphoinositide 3-kinase isoforms.
  JOURNAL   Cell. 2013 May 23;153(5):1050-63. doi: 10.1016/j.cell.2013.04.031.
  PUBMED    23706742
REFERENCE   2  (bases 1 to 5601)
  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: 
            "10.1016/j.cell.2013.04"; journalName: "Cell"; date: "2013-05-23- 
            23"; volume: "153"; issue: "5"; pages: "1050-63"
COMMENT     Sequence Label: RIT1
FEATURES             Location/Qualifiers
     source          1..5601
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..935
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(1651..1673)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1818..1836)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1904..2008
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2009..2077,2078..2869)
                     /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(2227..2246)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3040..3628
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3529..3548
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3782..3799
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3872..3949
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3950..5032
                     /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(3969..3988)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5081..5086,5087..5104,5105..5111)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5119..5135
                     /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     5143..5159
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5143..5159
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5155..5601,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5175..5192)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5175..5191)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5184..5206)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 agcttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gttccgcgtg gatctgagtc cggagctcgc cccattggta
      961 gcagctgtag cagccctgca gcactctccc gggaatacaa actagtgatg ctgggtgccg
     1021 gtggagttgg gaagagtgcc atgacaatgc agttcatcag ccaccgattc ccagaagacc
     1081 atgaccccac cattgaagat gcttataaga tccggatccg cattgatgat gaacctgcca
     1141 atctggacat cctggacaca gcaggacagg cagagtttac agccatgcgg gatcagtata
     1201 tgagggcagg agaaggattc atcatctgtt actccatcac ggatcgtcga agtttccacg
     1261 aagttcggga gtttaaacaa ctgatttacc gggtccgacg caccgatgat acaccggtgg
     1321 ttcttgtggg gaacaagtct gacctaaagc agctgaggca ggtctccaag gaagagggat
     1381 tgtctctggc tcgagaattc agttgtccct tttttgagac ctcagctgcc taccgttact
     1441 acatcgacga cgttttccac gccctcgtcc gggagatccg taagaaagag aaggagctag
     1501 tactggccat ggagaagaag gccaaaccca agaacagcgt atggaagagg ctgaagtcac
     1561 cgttcaggag gaagaaagac tcggtcacct gacaattcat cgtgactgac tgacgatctg
     1621 cctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt
     1681 cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg
     1741 tgttggcggg tgtcggggcg cagccatgac ccagtcacgt agcgatagcg gagtgtataa
     1801 ttcttgaaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     1861 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     1921 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     1981 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     2041 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     2101 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     2161 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     2221 agttctgcta tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg
     2281 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     2341 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     2401 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     2461 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     2521 accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact
     2581 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     2641 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     2701 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     2761 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     2821 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     2881 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     2941 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     3001 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     3061 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     3121 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     3181 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     3241 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     3301 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     3361 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     3421 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     3481 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     3541 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     3601 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     3661 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     3721 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     3781 cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca
     3841 tctgtgcggt atttcacacc gcataaattc cgacaccatc gaatggcgca aaacctttcg
     3901 cggtatggca tgatagcgcc cggaagagag tcaattcagg gtggtgaatg tgaaaccagt
     3961 aacgttatac gatgtcgcag agtatgccgg tgtctcttat cagaccgttt cccgcgtggt
     4021 gaaccaggcc agccacgttt ctgcgaaaac gcgggaaaaa gtggaagcgg cgatggcgga
     4081 gctgaattac attcccaacc gcgtggcaca acaactggcg ggcaaacagt cgttgctgat
     4141 tggcgttgcc acctccagtc tggccctgca cgcgccgtcg caaattgtcg cggcgattaa
     4201 atctcgcgcc gatcaactgg gtgccagcgt ggtggtgtcg atggtagaac gaagcggcgt
     4261 cgaagcctgt aaagcggcgg tgcacaatct tctcgcgcaa cgcgtcagtg ggctgatcat
     4321 taactatccg ctggatgacc aggatgccat tgctgtggaa gctgcctgca ctaatgttcc
     4381 ggcgttattt cttgatgtct ctgaccagac acccatcaac agtattattt tctcccatga
     4441 agacggtacg cgactgggcg tggagcatct ggtcgcattg ggtcaccagc aaatcgcgct
     4501 gttagcgggc ccattaagtt ctgtctcggc gcgtctgcgt ctggctggct ggcataaata
     4561 tctcactcgc aatcaaattc agccgatagc ggaacgggaa ggcgactgga gtgccatgtc
     4621 cggttttcaa caaaccatgc aaatgctgaa tgagggcatc gttcccactg cgatgctggt
     4681 tgccaacgat cagatggcgc tgggcgcaat gcgcgccatt accgagtccg ggctgcgcgt
     4741 tggtgcggat atctcggtag tgggatacga cgataccgaa gacagctcat gttatatccc
     4801 gccgttaacc accatcaaac aggattttcg cctgctgggg caaaccagcg tggaccgctt
     4861 gctgcaactc tctcagggcc aggcggtgaa gggcaatcag ctgttgcccg tctcactggt
     4921 gaaaagaaaa accaccctgg cgcccaatac gcaaaccgcc tctccccgcg cgttggccga
     4981 ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg
     5041 caattaatgt aagttagctc actcattagg caccccaggc tttacacttt atgcttccgg
     5101 ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc
     5161 atgattacgg attcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
     5221 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
     5281 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg gcgctttgcc
     5341 tggtttccgg caccagaagc ggtgccggaa agctggctgg agtgcgatct tcctgaggcc
     5401 gatactgtcg tcgtcccctc aaactggcag atgcacggtt acgatgcgcc catctacacc
     5461 aacgtgacct atcccattac ggtcaatccg ccgtttgttc ccacggagaa tccgacgggt
     5521 tgttactcgc tcacatttaa tgttgatgaa agctggctac aggaaggcca gacgcgaatt
     5581 atttttgatg gcgttggaat t
//
