LOCUS       sequence-226652-        8035 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express mutant RGS7- E383K- in mamalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8035)
  AUTHORS   Qutob N, Masuho I, Alon M, Emmanuel R, Cohen I, Di Pizio A, Madore 
            J, Elkahloun A, Ziv T, Levy R, Gartner JJ, Hill VK, Lin JC, Hevroni 
            Y, Greenberg P, Brodezki A, Rosenberg SA, Kosloff M, Hayward NK, 
            Admon A, Niv MY, Scolyer RA, Martemyanov KA, Samuels Y
  TITLE     RGS7 is recurrently mutated in melanoma and promotes migration and 
            invasion of human cancer cells.
  JOURNAL   Sci Rep. 2018 Jan 12;8(1):653. doi: 10.1038/s41598-017-18851-4.
  PUBMED    29330521
REFERENCE   2  (bases 1 to 8035)
  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: "Sci Rep."; 
            date: "2018-01-12"; volume: "8(1):653. doi"; pages: " 
            10.1038/s41598-017-18851-4"
COMMENT     Sequence Label: pCDF1-MCS2-EF1-Puro-RGS7-E383K
FEATURES             Location/Qualifiers
     source          1..8035
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..204
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     promoter        1347..1550
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1547..1571
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1640..1649
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3053..3076
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        3121..3332
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             3345..3613
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     CDS             3640..4239
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3640..3659)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4136..4156
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    4246..4834
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4299..4319)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4717..4728)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    5171..5292
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5208..5227)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5262..5281
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5332..5467
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5500..5516)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5500..5516)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5513..5535)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5524..5540
                     /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)."
     promoter        complement(join(5548..5554,5555..5572,5573..5578))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5593..5614
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5731..5748)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5902..6490)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5982..6001)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6661..7452,7453..7521))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7284..7303
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7522..7626)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7694..7712
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7750..7772)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7872..7891
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
       61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      181 tgggaggtct atataagcag agcttgtgaa acttcgagga gtctctttgt tgaggacttt
      241 tgagttctcc cttgaggctc ccacagatac aataaatatt tgagattgaa ccctgtcgag
      301 tatctgtgta atctttttta cctgtgaggt ctcggaatcc gggccgagaa cttcgcagtt
      361 ggcgcccgaa cagggacttg attgagagtg attgaggaag tgaagctaga gcaatagaaa
      421 gctgttaagc agaactcctg ctgacctaaa tagggaagca gtagcagacg ctgctaacag
      481 tgagtatctc tagtgaagca gactcgagct cataatcaag tcattgttta aaggcccaga
      541 taaattacat ctggtgactc ttcgcggacc ttcaagccag gagattcgcc gagggacagt
      601 caacaaggta ggagagattc tacagcaaca tggggaatgg acaggggcga gattggaaaa
      661 tggccattaa gagatgtagt aatgttgctg taggagtagg ggggaagagt aaaaaatttg
      721 gagaagggaa tttcagatgg gccattagaa tggctaatgt atctacagga cgagaacctg
      781 gtgatatacc agagacttta gatcaactaa ggttggttat ttgcgattta caagaaagaa
      841 gagaaaaatt tggatctagc aaagaaattg atatggcaat tcctgcattg aggagaaatg
      901 gtaggcaatg tggcatgtct gaaaaagagg aggaatgatg aagtatctca gacttatttt
      961 ataagggaga tactgtgctg agttcttccc tttgaggaag gtatgtcata tcctagacat
     1021 agtctcaatt ttaaaagaag aggtaggata ggagggatgg ccccttatga attattagca
     1081 caacaagaat ccttaagaat acaagattat ttttctgcaa taccacaaaa attgcaagca
     1141 cagtggattt attataaaga tcaaaaagat aagaaatgga aaggaccaat gagagtagaa
     1201 tactggggac agggatcagt attattaaag gatgaagaga agggatattt tcttataatc
     1261 gatactagta ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
     1321 acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg
     1381 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
     1441 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
     1501 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct cgtttagtga
     1561 accgtcagat cgcctggaga cgccatccac gctgttttga cctccataga agattctaga
     1621 gcccgggcgc gccggatccg ccaccatggc ccaggggaat aattatgggc agaccagcaa
     1681 cggggtggcc gatgaatcac ccaacatgct ggtgtacaga aagatggaag acgtcatagc
     1741 acggatgcaa gatgaaaaaa atggaattcc tattcgtacg gtcaaaagct ttctttccaa
     1801 gatacctagc gtcttctctg gttcagacat tgttcaatgg ttgataaaga acttaactat
     1861 agaagatcca gtggaggcgc tccatttggg aacattaatg gctgcccacg gctacttctt
     1921 tccaatctca gatcatgtcc tcacactcaa ggatgatggc accttttacc ggtttcaaac
     1981 cccctatttt tggccatcaa attgttggga gccggaaaac acagattatg ccgtttacct
     2041 ctgcaagaga acaatgcaaa acaaggcacg actggagctc gcagactatg aggctgagag
     2101 cctggccagg ctgcagagag catttgcccg gaagtgggag ttcattttca tgcaagcaga
     2161 agcacaagca aaagtggaca agaagagaga caagattgaa aggaagatcc ttgacagcca
     2221 agagagagcg ttctgggacg tgcacaggcc cgtgcctgga tgtgtaaata caactgaagt
     2281 ggacattaag aagtcatcca gaatgagaaa cccccacaaa acacggaagt ctgtctatgg
     2341 tttacaaaat gatattagaa gtcacagtcc tacccacaca cccacaccag aaactaaacc
     2401 tccaacagaa gatgagttac aacaacagat aaaatattgg caaatacagt tagatagaca
     2461 tcggttaaaa atgtcaaaag tcgctgacag tctactaagt tacacggaac agtatttaga
     2521 atacgacccg tttcttttgc cacctgaccc ttctaaccca tggctgtccg atgacaccac
     2581 tttctgggaa cttgaggcaa gcaaagaacc gagccagcag agggtaaaac gatggggttt
     2641 tggcatggac gaggcattga aagacccagt tgggagagaa cagttcctta aatttctaga
     2701 gtcagaattc agctcggaaa atttaagatt ctggctggca gtggaggacc tgaaaaagag
     2761 gcctattaaa gaagtaccct caagagttca gaaaatatgg caagagtttc tggctcccgg
     2821 agcccccagt gctattaact tggattccaa gagttatgac aaaaccacac agaacgtgaa
     2881 ggaacctgga cgatacacat ttgaagatgc tcaggagcac atttacaaac tgatgaaaag
     2941 tgattcatac ccacgtttta taagatccag tgcctatcag gagcttctac aggcaaagaa
     3001 aaaggggaaa tctctcacgt ccaagaggtt aacaagcctt gctcagtctt acgattacaa
     3061 ggatgacgac gataagtaag cggccgcgaa ggatctgcga tcgctccggt gcccgtcagt
     3121 gggcagagcg cacatcgccc acagtccccg agaagttggg gggaggggtc ggcaattgaa
     3181 cgggtgccta gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg tactggctcc
     3241 gcctttttcc cgagggtggg ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc
     3301 tttttcgcaa cgggtttgcc gccagaacac agctgaagct tcgaggggct cgcatctctc
     3361 cttcacgcgc ccgccgccct acctgaggcc gccatccacg ccggttgagt cgcgttctgc
     3421 cgcctcccgc ctgtggtgcc tcctgaactg cgtccgccgt ctaggtaagt ttaaagctca
     3481 ggtcgagacc gggcctttgt ccggcgctcc cttggagcct acctagactc agccggctct
     3541 ccacgctttg cctgaccctg cttgctcaac tctacgtctt tgtttcgttt tctgttctgc
     3601 gccgttacag atccaagctg tgaccggcgc ctacgctaga tgaccgagta caagcccacg
     3661 gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc
     3721 gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc
     3781 gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg
     3841 gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg
     3901 ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa
     3961 cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc
     4021 gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga
     4081 gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac
     4141 ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga
     4201 ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgag tcgacaatca acctctggat
     4261 tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt
     4321 ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc
     4381 tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg
     4441 caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc
     4501 accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa
     4561 ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat
     4621 tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc
     4681 tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt
     4741 ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag
     4801 acgagtcgga tctccctttg ggccgcctcc ccgcctggta ccgatgacag agttagaaga
     4861 tcgcttcagg aagctatttg gcacgacttc tacaacggga gacagcacag tagattctga
     4921 agatgaacct cctaaaaaag aaaaaagggt ggactgggat gagtattgga accctgaaat
     4981 cgatagcttc cagtgctttg tgaaacttcg aggagtctct ttgttgagga cttttgagtt
     5041 ctcccttgag gctcccacag atacaataaa tatttgagat tgaaccctgt cgagtatctg
     5101 tgtaatcttt tttacctgtg aggtctcgga atccgggccg agaacttcgc agcgagctca
     5161 ttgtaccgcg aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     5221 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     5281 caatgtatct tatcatgtct ggctctagct atcccgcccc taactccgcc catcccgccc
     5341 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     5401 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     5461 ggaggcctag acttttgcag agaccaaatt cgtaatcatg tcatagctgt ttcctgtgtg
     5521 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     5581 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     5641 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     5701 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     5761 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     5821 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     5881 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     5941 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     6001 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     6061 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     6121 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     6181 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     6241 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     6301 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     6361 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     6421 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     6481 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     6541 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     6601 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     6661 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     6721 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     6781 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     6841 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     6901 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     6961 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     7021 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     7081 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     7141 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     7201 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     7261 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     7321 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     7381 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     7441 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     7501 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     7561 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     7621 tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac
     7681 attaacctat aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga
     7741 cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga
     7801 tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg
     7861 gcttaactat gcggcatcag agcagattgt actgagagtg caccatatgc caagtacgcc
     7921 ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt
     7981 atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta ccatg
//
