LOCUS       sequence-63776-e        6441 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses rM3D (Gs) neuronal activator.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6441)
  AUTHORS   Armbruster BN, Li X, Pausch MH, Herlitze S, Roth BL
  TITLE     Evolving the lock to fit the key to create a family of G 
            protein-coupled receptors potently activated by an inert ligand.
  JOURNAL   Proc Natl Acad Sci U S A. 2007 Mar 20;104(12):5163-8. Epub 2007 Mar 
            2.
  PUBMED    17360345
REFERENCE   2  (bases 1 to 6441)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2007-03-20"; volume: "104(12)"; pages: 
            "5163-8. Epub 2007 Mar 2"
COMMENT     Sequence Label: pcDNA5/FRT-HA-rM3D(Gs)
FEATURES             Location/Qualifiers
     source          1..6441
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1010..1036
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(2394..2411)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2400..2624
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2757..2776)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2907..2954
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             2962..3984
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2962..3984
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    4114..4235
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4151..4170)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4205..4224
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4284..4300)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4284..4300)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4297..4319)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4308..4324
                     /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(4332..4338,4339..4356,4357..4362))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4377..4398
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4515..4532)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4686..5274)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4766..4785)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5445..6236,6237..6305))
                     /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     6068..6087
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6306..6410)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattctgc
      961 agataagctg ctgcaggtcg tcctgggact atgtcagaga gtcacaatgt acccctacga
     1021 cgtccccgac tacgccacct tgcacagtaa cagtacaacc tcgcctttgt ttcccaacat
     1081 cagctcttcc tgggtgcaca gtccctcgga ggcagggctg cccttgggga cagtcactca
     1141 gttgggcagc tacaacattt cacaagaaac tgggaatttc tcctcaaacg acacctccag
     1201 cgaccctctc gggggtcaca ccatctggca agtggtcttc attgccttct taactggctt
     1261 cctggcattg gtgaccatca ttggcaacat ccttgtcatt gtggccttca aggtcaacaa
     1321 acagctgaag acagtcaaca actacttcct cttaagcctg gcctgtgcag acctgatcat
     1381 cggggtcatt tccatgaacc tgttcactac ctacatcatt atgaaccgtt gggcactggg
     1441 gaacttagcc tgcgacctct ggctctccat tgactgtgtg gccagcaatg cctctgtcat
     1501 gaatctgctg gtcatcagct ttgacaggta cttttccatc acttctccat tccgctacca
     1561 gagcctgatg accagggctc gagctggtgt gatgattggt ctggcttggg tcatctcctt
     1621 tgtcctatgg gctcctgcca tcttgttctg gcaatacttt gtagggaaga gaactgtgcc
     1681 cccaggagaa tgtttcattc agtttctgag tgagcccacc atcaccttcg gcacggcgat
     1741 cgctggcttt tacatgcctg tcaccatcat gactatttta tactggcggg tgtaccggga
     1801 ggccaaggag cagatcagga agatcgaccg ctgcgagggc cggttctatg gcagccagga
     1861 gcagccgcag ccacccccgc tcccccaaca ccagcccatc ctcggcaacg gccgtgccag
     1921 caagaggaag acgtcccgtg tcatggccat gagggaacac aaagctctgc agacgctcag
     1981 tgccatcttg ctagccttca tcatcacgtg gaccccctac aacatcatgg tcctggtgaa
     2041 caccttctgt gacagctgca tacccaaaac ctattggaat ctgggctact ggctgtgcta
     2101 tatcaacagc accgtgaacc ctgtgtgcta tgccctgtgc aacaaaacat tcagaaccac
     2161 cttcaagacg ctcctcttgt gccagtgtga caaaaggaag aggcgcaaac agcagtacca
     2221 gcagagacag tcggtcattt ttcacaagcg agtgccggag caggccttgt agaaaagggg
     2281 tatgttgaga gcagtgacca cgcaagcgcg tcagcccaca cagcctagca ggagtcatcc
     2341 agcacagtgg cggccgctcg agtctagagg gcccgtttaa acccgctgat cagcctcgac
     2401 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     2461 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     2521 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     2581 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag
     2641 aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc
     2701 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     2761 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     2821 tcgggggtcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     2881 gattagggtg atggttcacg tacctagaag ttcctattcc gaagttccta ttctctagaa
     2941 agtataggaa cttccttggc caaaaagcct gaactcaccg cgacgtctgt cgagaagttt
     3001 ctgatcgaaa agttcgacag cgtctccgac ctgatgcagc tctcggaggg cgaagaatct
     3061 cgtgctttca gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa tagctgcgcc
     3121 gatggtttct acaaagatcg ttatgtttat cggcactttg catcggccgc gctcccgatt
     3181 ccggaagtgc ttgacattgg ggaattcagc gagagcctga cctattgcat ctcccgccgt
     3241 gcacagggtg tcacgttgca agacctgcct gaaaccgaac tgcccgctgt tctgcagccg
     3301 gtcgcggagg ccatggatgc gatcgctgcg gccgatctta gccagacgag cgggttcggc
     3361 ccattcggac cgcaaggaat cggtcaatac actacatggc gtgatttcat atgcgcgatt
     3421 gctgatcccc atgtgtatca ctggcaaact gtgatggacg acaccgtcag tgcgtccgtc
     3481 gcgcaggctc tcgatgagct gatgctttgg gccgaggact gccccgaagt ccggcacctc
     3541 gtgcacgcgg atttcggctc caacaatgtc ctgacggaca atggccgcat aacagcggtc
     3601 attgactgga gcgaggcgat gttcggggat tcccaatacg aggtcgccaa catcttcttc
     3661 tggaggccgt ggttggcttg tatggagcag cagacgcgct acttcgagcg gaggcatccg
     3721 gagcttgcag gatcgccgcg gctccgggcg tatatgctcc gcattggtct tgaccaactc
     3781 tatcagagct tggttgacgg caatttcgat gatgcagctt gggcgcaggg tcgatgcgac
     3841 gcaatcgtcc gatccggagc cgggactgtc gggcgtacac aaatcgcccg cagaagcgcg
     3901 gccgtctgga ccgatggctg tgtagaagta ctcgccgata gtggaaaccg acgccccagc
     3961 actcgtccga gggcaaagga atagcacgta ctacgagatt tcgattccac cgccgccttc
     4021 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     4081 ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt
     4141 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     4201 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct
     4261 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     4321 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     4381 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     4441 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     4501 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     4561 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     4621 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     4681 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     4741 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     4801 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     4861 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     4921 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     4981 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     5041 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     5101 tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca
     5161 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     5221 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     5281 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     5341 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     5401 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     5461 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     5521 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     5581 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     5641 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     5701 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     5761 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     5821 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     5881 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     5941 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     6001 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     6061 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     6121 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     6181 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     6241 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     6301 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     6361 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     6421 cgaaaagtgc cacctgacgt c
//
