LOCUS       sequence-483123-        5717 bp    DNA     circular SYN 19-AUG-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 5717)
  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 5717)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 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..5717
                     /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    1300..1324
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(1977..1979,1980..1982,1983..2696)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2022..2043)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2283..2302)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2630..2651
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(2726..2750)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2776..2792)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2776..2792)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(3529..3549)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3529..3547)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3568..3584)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3568..3584)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3581..3603)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3592..3608
                     /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(3616..3622,3623..3640,3641..3646))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3661..3682
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3799..3816)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3970..4558)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4050..4069)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4729..5520,5521..5589))
                     /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     5352..5371
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5590..5694)
                     /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 tctagcgcca
      781 aggcaatctt cgtaaaaaat aacaatagag tcatccctca gaacagaaaa gaaacccgcc
      841 cgggggctca aatacacatg tcatttttct tttttcgcct gatttttagc ctaaattagt
      901 gtttagcaat tttttaaggt aactcggtgg agtcatttga actttttaaa ctaaaatttc
      961 agaaaaaatt cttttttttt gtcaaaaaat aattttcccg ccggaatttt tcaaaaaaat
     1021 aatttttatt ttatttttca agttccttgt ttcctcttta tttttttatt ttgcccccaa
     1081 aagttttaca ctaaaatatt taattttcag gtaattcatc gaaagtcttt ctattttccg
     1141 catctcttgt tcaaaagctt gcatgcctgc aggtcgactc tagaggatcc ccgggattgg
     1201 ccaaaggacc caaaggtatg tttcgaatga tactaacata acatagaaca ttttcaggag
     1261 gacccttgga gggtaccggt agaaaaaaag cttgatatca caagtttgta caaaaaagca
     1321 ggctccgaat tcgcccttgg actagtcata tgggtaccat gcaagcaatc aaatgtgtcg
     1381 tcgttggtga cgtagccgtc ggtaagacgt gtctcctgct atcgtacact acaaacgctt
     1441 ttcccggaga atatattcta acggtattcg acacctactc aacaaatgtg atggtcgacg
     1501 gaaggccaat aaatctcagc ctatgggaca cagctggaca ggacgattac gatcaattcc
     1561 gccacctgtc atttccacaa acagacgtat tcctcgtatg ctttgcactg aataatccag
     1621 caagttttga gaatgttcgt gcaaaatggt atccagaagt atcacatcat tgcccaaata
     1681 caccgattat tttggttgga actaaagctg atttacgcga ggatcgagat actattgaac
     1741 ggctccgtga acgtcggctc caaccagtga gccacaccca gggttacgtg atggcaaagg
     1801 aaatcaaggc ggtcaagtac ctggaatgct cggcgcttac ccaaattgga ttgaaacaag
     1861 ttttcgatga ggcaattcgt actgggctca ccccgccaca aacaccacaa acgagagcca
     1921 aaaagagcaa ttgcacggtg ggtaccagtg gttccggagg atctggagga tctggtatgg
     1981 tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg
     2041 acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca
     2101 agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg
     2161 tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcagc
     2221 acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca
     2281 aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga
     2341 accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc
     2401 tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca
     2461 tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc
     2521 actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc
     2581 tgagcaccca gtccaagctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc
     2641 tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagtagcata
     2701 tggcggccgc ggaagggcga attcgaccca gctttcttgt acaaagtggt gatatcgaat
     2761 tcctgcagcc cgggggatcc actagttcta gagcggccgc caccgcggcc gctgtcatca
     2821 gatcgccatc tcgcgcccgt gcctctgact tctaagtcca attactcttc aacatcccta
     2881 catgctcttt ctccctgtgc tcccaccccc tatttttgtt attatcaaaa aacttctctt
     2941 aatttctttg ttttttagct tcttttaagt cacctctaac aatgaaattg tgtagattca
     3001 aaaatagaat taattcgtaa taaaaagtcg aaaaaaattg tgctccctcc ccccattaat
     3061 aataattcta tcccaaaatc tacacaatgt tctgtgtaca cttcttatgt tttttacttc
     3121 tgataaattt ttttgaaaca tcatagaaaa aaccgcacac aaaatacctt atcatatgtt
     3181 acgtttcagt ttatgaccgc aatttttatt tcttcgcacg tctgggcctc tcatgacgtc
     3241 aaatcatgct catcgtgaaa aagttttgga gtatttttgg aatttttcaa tcaagtgaaa
     3301 gtttatgaaa ttaattttcc tgcttttgct ttttggggtt tcccctattg tttgtcaaga
     3361 tttcgaggac ggcgtttttc ttgctaaaat cacaagtatt gatgagcacg atgcaagaaa
     3421 gatcggaaga aggtttgggt ttgaggctca gtggaaggtg agtagaagtt gataatttga
     3481 aagtggagta gtgtctatgg ggtttttgcc gagctccagc ttttgttccc tttagtgagg
     3541 gttaattgcg cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     3601 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     3661 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     3721 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     3781 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     3841 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     3901 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     3961 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     4021 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     4081 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     4141 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     4201 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     4261 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     4321 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     4381 gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa
     4441 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     4501 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     4561 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     4621 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     4681 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     4741 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     4801 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     4861 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     4921 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     4981 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     5041 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     5101 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     5161 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     5221 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     5281 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     5341 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     5401 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     5461 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     5521 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     5581 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     5641 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     5701 tccccgaaaa gtgccac
//
