LOCUS       sequence-221944-        7430 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Express double mutant RGS7- R44C+V65C in mamalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 7430)
  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 7430)
  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-R44C+V56C
FEATURES             Location/Qualifiers
     source          1..7430
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     polyA_signal    132..266
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(476..1064)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(1235..2026,2027..2095))
                     /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     1858..1877
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2096..2200)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2268..2286
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2324..2346)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2446..2465
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        2610..2813
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2764..2784
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2810..2834
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      2903..2912
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4316..4339
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        4384..4595
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             4608..4876
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     CDS             4903..5502
                     /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(4903..4922)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5399..5419
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    5509..6097
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5562..5582)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5980..5991)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    6434..6555
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      6595..6730
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6763..6779)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6763..6779)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6776..6798)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6787..6803
                     /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(6811..6817,6818..6835,6836..6841))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6856..6877
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     misc_feature    7396..7414
                     /label=Tn5 ME
                     /note="hyperactive mosaic end for Tn5 transposase 
                     recognition (Reznikoff et al., 2004)"
ORIGIN
        1 cagcccttga ggctcccaca gatacaataa atatttgaga ttgaaccctg tcgagtatct
       61 gtgtaatctt ttttacctgt gaggtctcgg aatccgggcc gagaacttcg cagcgagctc
      121 attgtaccgc gaacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
      181 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
      241 tcaatgtatc ttatcatgtc tggatccccg ggtaccgagc tcgaattaak ysctcttccg
      301 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
      361 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
      421 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
      481 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
      541 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
      601 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
      661 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
      721 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
      781 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
      841 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
      901 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
      961 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     1021 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     1081 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     1141 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     1201 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     1261 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     1321 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     1381 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     1441 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     1501 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     1561 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     1621 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     1681 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     1741 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     1801 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     1861 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     1921 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     1981 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     2041 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     2101 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     2161 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     2221 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca
     2281 cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc
     2341 tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg
     2401 gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga
     2461 ttgtactgag agtgcaccat atgccaagta cgccccctat tgacgtcaat gacggtaaat
     2521 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
     2581 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
     2641 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
     2701 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
     2761 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctcgtttag
     2821 tgaaccgtca gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagattct
     2881 agagcccggg cgcgccggat ccgccaccat ggcccagggg aataattatg ggcagaccag
     2941 caacggggtg gccgatgaat cacccaacat gctggtgtac agaaagatgg aagacgtcat
     3001 agcacggatg caagatgaaa aaaatggaat tcctatttgt acggtcaaaa gctttctttc
     3061 caagatacct agctgcttct ctggttcaga cattgttcaa tggttgataa agaacttaac
     3121 tatagaagat ccagtggagg cgctccattt gggaacatta atggctgccc acggctactt
     3181 ctttccaatc tcagatcatg tcctcacact caaggatgat ggcacctttt accggtttca
     3241 aaccccctat ttttggccat caaattgttg ggagccggaa aacacagatt atgccgttta
     3301 cctctgcaag agaacaatgc aaaacaaggc acgactggag ctcgcagact atgaggctga
     3361 gagcctggcc aggctgcaga gagcatttgc ccggaagtgg gagttcattt tcatgcaagc
     3421 agaagcacaa gcaaaagtgg acaagaagag agacaagatt gaaaggaaga tccttgacag
     3481 ccaagagaga gcgttctggg acgtgcacag gcccgtgcct ggatgtgtaa atacaactga
     3541 agtggacatt aagaagtcat ccagaatgag aaacccccac aaaacacgga agtctgtcta
     3601 tggtttacaa aatgatatta gaagtcacag tcctacccac acacccacac cagaaactaa
     3661 acctccaaca gaagatgagt tacaacaaca gataaaatat tggcaaatac agttagatag
     3721 acatcggtta aaaatgtcaa aagtcgctga cagtctacta agttacacgg aacagtattt
     3781 agaatacgac ccgtttcttt tgccacctga cccttctaac ccatggctgt ccgatgacac
     3841 cactttctgg gaacttgagg caagcaaaga accgagccag cagagggtaa aacgatgggg
     3901 ttttggcatg gacgaggcat tgaaagaccc agttgggaga gaacagttcc ttaaatttct
     3961 agagtcagaa ttcagctcgg aaaatttaag attctggctg gcagtggagg acctgaaaaa
     4021 gaggcctatt aaagaagtac cctcaagagt tcaggaaata tggcaagagt ttctggctcc
     4081 cggagccccc agtgctatta acttggattc caagagttat gacaaaacca cacagaacgt
     4141 gaaggaacct ggacgataca catttgaaga tgctcaggag cacatttaca aactgatgaa
     4201 aagtgattca tacccacgtt ttataagatc cagtgcctat caggagcttc tacaggcaaa
     4261 gaaaaagggg aaatctctca cgtccaagag gttaacaagc cttgctcagt cttacgatta
     4321 caaggatgac gacgataagt aagcggccgc gaaggatctg cgatcgctcc ggtgcccgtc
     4381 agtgggcaga gcgcacatcg cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt
     4441 gaacgggtgc ctagagaagg tggcgcgggg taaactggga aagtgatgtc gtgtactggc
     4501 tccgcctttt tcccgagggt gggggagaac cgtatataag tgcagtagtc gccgtgaacg
     4561 ttctttttcg caacgggttt gccgccagaa cacagctgaa gcttcgaggg gctcgcatct
     4621 ctccttcacg cgcccgccgc cctacctgag gccgccatcc acgccggttg agtcgcgttc
     4681 tgccgcctcc cgcctgtggt gcctcctgaa ctgcgtccgc cgtctaggta agtttaaagc
     4741 tcaggtcgag accgggcctt tgtccggcgc tcccttggag cctacctaga ctcagccggc
     4801 tctccacgct ttgcctgacc ctgcttgctc aactctacgt ctttgtttcg ttttctgttc
     4861 tgcgccgtta cagatccaag ctgtgaccgg cgcctacgct agatgaccga gtacaagccc
     4921 acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct cgccgccgcg
     4981 ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat cgagcgggtc
     5041 accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc
     5101 gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga agcgggggcg
     5161 gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct ggccgcgcag
     5221 caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc
     5281 accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc
     5341 ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacctc cgcgccccgc
     5401 aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa
     5461 ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gagtcgacaa tcaacctctg
     5521 gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta
     5581 tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt
     5641 ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc
     5701 aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt
     5761 gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg
     5821 gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac
     5881 aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc
     5941 acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac
     6001 cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct
     6061 cagacgagtc ggatctccct ttgggccgcc tccccgcctg gtaccgatga cagagttaga
     6121 agatcgcttc aggaagctat ttggcacgac ttctacaacg ggagacagca cagtagattc
     6181 tgaagatgaa cctcctaaaa aagaaaaaag ggtggactgg gatgagtatt ggaaccctga
     6241 aatcgatagc ttccagtgct ttgtgaaact tcgaggagtc tctttgttga ggacttttga
     6301 gttctccctt gaggctccca cagatacaat aaatatttga gattgaaccc tgtcgagtat
     6361 ctgtgtaatc ttttttacct gtgaggtctc ggaatccggg ccgagaactt cgcagcgagc
     6421 tcattgtacc gcgaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     6481 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     6541 catcaatgta tcttatcatg tctggctcta gctatcccgc ccctaactcc gcccatcccg
     6601 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     6661 tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt
     6721 tttggaggcc tagacttttg cagagaccaa attcgtaatc atgtcatagc tgtttcctgt
     6781 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     6841 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     6901 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     6961 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     7021 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     7081 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     7141 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     7201 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     7261 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     7321 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     7381 cagttcggtg taggtctgtc tcttatacac atctccgagc ccacgagacc
//
