LOCUS       sequence-486783-        5710 bp    DNA     circular SYN 19-SEP-2025
DEFINITION  Expresses rac-2(G12V)::mGFP in cholinergic motor neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5710)
  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 5710)
  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: pSJC321
FEATURES             Location/Qualifiers
     source          1..5710
                     /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(1970..1972,1973..1975,1976..2689)
                     /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(2015..2036)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2276..2295)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2623..2644
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(2719..2743)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2769..2785)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2769..2785)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(3522..3542)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3522..3540)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3561..3577)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3561..3577)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3574..3596)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3585..3601
                     /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(3609..3615,3616..3633,3634..3639))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3654..3675
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3792..3809)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3963..4551)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4043..4062)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4722..5513,5514..5582))
                     /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     5345..5364
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5583..5687)
                     /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 catgcaagca atcaaatgtg tcgtcgttgg
     1381 tgacgtagcc gtcggtaaga cgtgtctcct gctatcgtac actacaaacg cttttcccgg
     1441 agaatatatt ctaacggtat tcgacaccta ctcaacaaat gtgatggtcg acggaaggcc
     1501 aataaatctc agcctatggg acacagctgg acaggacgat tacgatcaat tccgccacct
     1561 gtcatttcca caaacagacg tattcctcgt atgctttgca ctgaataatc cagcaagttt
     1621 tgagaatgtt cgtgcaaaat ggtatccaga agtatcacat cattgcccaa atacaccgat
     1681 tattttggtt ggaactaaag ctgatttacg cgaggatcga gatactattg aacggctccg
     1741 tgaacgtcgg ctccaaccag tgagccacac ccagggttac gtgatggcaa aggaaatcaa
     1801 ggcggtcaag tacctggaat gctcggcgct tacccaaatt ggattgaaac aagttttcga
     1861 tgaggcaatt cgtactgggc tcaccccgcc acaaacacca caaacgagag ccaaaaagag
     1921 caattgcacg gtgggtacca gtggttccgg aggatctgga ggatctggta tggtgagcaa
     1981 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
     2041 cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac
     2101 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
     2161 cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt
     2221 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga
     2281 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
     2341 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
     2401 caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt
     2461 gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca
     2521 gcagaacacc cccatcggcg acggccccgt gctgctgtcc gacaaccact acctgagcac
     2581 ccagtccaag ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
     2641 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtagc atatggcggc
     2701 cgcggaaggg cgaattcgac ccagctttct tgtacaaagt ggtgatatcg aattcctgca
     2761 gcccggggga tccactagtt ctagagcggc cgccaccgcg gccgctgtca tcagatcgcc
     2821 atctcgcgcc cgtgcctctg acttctaagt ccaattactc ttcaacatcc ctacatgctc
     2881 tttctccctg tgctcccacc ccctattttt gttattatca aaaaacttct cttaatttct
     2941 ttgtttttta gcttctttta agtcacctct aacaatgaaa ttgtgtagat tcaaaaatag
     3001 aattaattcg taataaaaag tcgaaaaaaa ttgtgctccc tccccccatt aataataatt
     3061 ctatcccaaa atctacacaa tgttctgtgt acacttctta tgttttttac ttctgataaa
     3121 tttttttgaa acatcataga aaaaaccgca cacaaaatac cttatcatat gttacgtttc
     3181 agtttatgac cgcaattttt atttcttcgc acgtctgggc ctctcatgac gtcaaatcat
     3241 gctcatcgtg aaaaagtttt ggagtatttt tggaattttt caatcaagtg aaagtttatg
     3301 aaattaattt tcctgctttt gctttttggg gtttccccta ttgtttgtca agatttcgag
     3361 gacggcgttt ttcttgctaa aatcacaagt attgatgagc acgatgcaag aaagatcgga
     3421 agaaggtttg ggtttgaggc tcagtggaag gtgagtagaa gttgataatt tgaaagtgga
     3481 gtagtgtcta tggggttttt gccgagctcc agcttttgtt ccctttagtg agggttaatt
     3541 gcgcgcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     3601 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     3661 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     3721 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     3781 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     3841 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     3901 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     3961 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     4021 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     4081 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     4141 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     4201 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     4261 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     4321 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     4381 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     4441 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     4501 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     4561 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     4621 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     4681 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     4741 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     4801 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     4861 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     4921 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     4981 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     5041 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     5101 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     5161 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     5221 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     5281 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     5341 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     5401 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     5461 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     5521 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     5581 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     5641 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     5701 aaagtgccac
//
