LOCUS       sequence-486786-        5692 bp    DNA     circular SYN 19-SEP-2025
DEFINITION  Expresses rap-1(G12V)::mGFP in cholinergic motor neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5692)
  AUTHORS   Lamb R, Scales M, Watkins J, Werner M, Cherra SJ
  TITLE     A PDZ-RapGEF promotes synaptic development in C. elegans through a 
            Rap/Rac signaling pathway.
  JOURNAL   Development. 2025 Aug 1:dev.204678. doi: 10.1242/dev.204678.
  PUBMED    40748210
REFERENCE   2  (bases 1 to 5692)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 19, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Development. 2025 Aug 1:dev.204678. doi: 10.1242/dev.204678."
COMMENT     Sequence Label: pSJC319
FEATURES             Location/Qualifiers
     source          1..5692
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1293..1317
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(1952..1954,1955..1957,1958..2671)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLSDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1997..2018)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2258..2277)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2605..2626
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(2701..2725)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2751..2767)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2751..2767)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(3504..3524)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3504..3522)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3543..3559)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3543..3559)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3556..3578)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3567..3583
                     /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(3591..3597,3598..3615,3616..3621))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3636..3657
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3774..3791)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3945..4533)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4025..4044)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4704..5495,5496..5564))
                     /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     5327..5346
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5565..5669)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tgggtaccgc
      661 ttttggtttt cacaattttc tggtttttgg gatttttgcg ggaattttga gaggagaaag
      721 ttgggagaga agatattgga atggtccccg gggagctgta catttataca ccaaggcaat
      781 cttcgtaaaa aataacaata gagtcatccc tcagaacaga aaagaaaccc gcccgggggc
      841 tcaaatacac atgtcatttt tcttttttcg cctgattttt agcctaaatt agtgtttagc
      901 aattttttaa ggtaactcgg tggagtcatt tgaacttttt aaactaaaat ttcagaaaaa
      961 attctttttt tttgtcaaaa aataattttc ccgccggaat ttttcaaaaa aataattttt
     1021 attttatttt tcaagttcct tgtttcctct ttattttttt attttgcccc caaaagtttt
     1081 acactaaaat atttaatttt caggtaattc atcgaaagtc tttctatttt ccgcatctct
     1141 tgttcaaaag cttgcatgcc tgcaggtcga ctctagagga tccccgggat tggccaaagg
     1201 acccaaaggt atgtttcgaa tgatactaac ataacataga acattttcag gaggaccctt
     1261 ggagggtacc ggtagaaaaa aagcttgata tcacaagttt gtacaaaaaa gcaggctccg
     1321 aattcgccct tggactagtc atatgggtac catgcgggag tataagattg ttgtgctcgg
     1381 atctgtcgga gtaggaaaat ccgcactgac ggtacaattc gtgcagggaa ttttcgtaga
     1441 gaagtatgac ccaacaattg aggatagtta cagaaagcaa gtggaagtcg acggccagca
     1501 atgtatgctt gagatactgg atacagcagg aacggagcaa ttcacagcga tgcgtgattt
     1561 gtacatgaag aatggacaag gatttgttct cgtttattca attactgctc aatcgacatt
     1621 caatgatctt atggatcttc gtgatcagat tcttcgtgtc aaagatacgg acgaggttcc
     1681 aatgattttg gtaggaaata agtgtgattt ggaagacgaa cgagttgttg gaaaagatca
     1741 gggacagaat ctcgccaggc agtttggatc agcattcctt gaaacctctg caaaagcaaa
     1801 gattaacgtc agcgaggtgt tctacgacct cgttcgtcaa ataaaccgac gttatcccga
     1861 atctggtcga cgccagggac aatcaaataa gcagtgctgc tcgtgtgtca tcatgggtac
     1921 cagtggttcc ggaggatctg gaggatctgg tatggtgagc aagggcgagg agctgttcac
     1981 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt
     2041 gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac
     2101 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca
     2161 gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
     2221 cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg
     2281 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga
     2341 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa
     2401 cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca
     2461 caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
     2521 cgacggcccc gtgctgctgt ccgacaacca ctacctgagc acccagtcca agctgagcaa
     2581 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat
     2641 cactctcggc atggacgagc tgtacaagta gcatatggcg gccgcggaag ggcgaattcg
     2701 acccagcttt cttgtacaaa gtggtgatat cgaattcctg cagcccgggg gatccactag
     2761 ttctagagcg gccgccaccg cggccgctgt catcagatcg ccatctcgcg cccgtgcctc
     2821 tgacttctaa gtccaattac tcttcaacat ccctacatgc tctttctccc tgtgctccca
     2881 ccccctattt ttgttattat caaaaaactt ctcttaattt ctttgttttt tagcttcttt
     2941 taagtcacct ctaacaatga aattgtgtag attcaaaaat agaattaatt cgtaataaaa
     3001 agtcgaaaaa aattgtgctc cctcccccca ttaataataa ttctatccca aaatctacac
     3061 aatgttctgt gtacacttct tatgtttttt acttctgata aatttttttg aaacatcata
     3121 gaaaaaaccg cacacaaaat accttatcat atgttacgtt tcagtttatg accgcaattt
     3181 ttatttcttc gcacgtctgg gcctctcatg acgtcaaatc atgctcatcg tgaaaaagtt
     3241 ttggagtatt tttggaattt ttcaatcaag tgaaagttta tgaaattaat tttcctgctt
     3301 ttgctttttg gggtttcccc tattgtttgt caagatttcg aggacggcgt ttttcttgct
     3361 aaaatcacaa gtattgatga gcacgatgca agaaagatcg gaagaaggtt tgggtttgag
     3421 gctcagtgga aggtgagtag aagttgataa tttgaaagtg gagtagtgtc tatggggttt
     3481 ttgccgagct ccagcttttg ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca
     3541 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     3601 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     3661 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     3721 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     3781 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     3841 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     3901 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     3961 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4021 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4081 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4141 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4201 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4261 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4321 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4381 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4441 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4501 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     4561 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     4621 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     4681 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     4741 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     4801 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     4861 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     4921 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     4981 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5041 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5101 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5161 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5221 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5281 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     5341 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     5401 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     5461 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     5521 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     5581 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     5641 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc ac
//
