LOCUS       sequence-483114-        5699 bp    DNA     circular SYN 19-AUG-2025
DEFINITION  Expresses rap-1::mGFP in cholinergic motor neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5699)
  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 5699)
  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: pBD31
FEATURES             Location/Qualifiers
     source          1..5699
                     /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(1959..1961,1962..1964,1965..2678)
                     /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(2004..2025)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2265..2284)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2612..2633
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(2708..2732)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2758..2774)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2758..2774)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(3511..3531)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3511..3529)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3550..3566)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3550..3566)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3563..3585)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3574..3590
                     /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(3598..3604,3605..3622,3623..3628))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3643..3664
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3781..3798)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3952..4540)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4032..4051)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4711..5502,5503..5571))
                     /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     5334..5353
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5572..5676)
                     /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 gcgggagtat aagattgttg
     1381 tgctcggatc tggaggagta ggaaaatccg cactgacggt acaattcgtg cagggaattt
     1441 tcgtagagaa gtatgaccca acaattgagg atagttacag aaagcaagtg gaagtcgacg
     1501 gccagcaatg tatgcttgag atactggata cagcaggaac ggagcaattc acagcgatgc
     1561 gtgatttgta catgaagaat ggacaaggat ttgttctcgt ttattcaatt actgctcaat
     1621 cgacattcaa tgatcttatg gatcttcgtg atcagattct tcgtgtcaaa gatacggacg
     1681 aggttccaat gattttggta ggaaataagt gtgatttgga agacgaacga gttgttggaa
     1741 aagatcaggg acagaatctc gccaggcagt ttggatcagc attccttgaa acctctgcaa
     1801 aagcaaagat taacgtcagc gaggtgttct acgacctcgt tcgtcaaata aaccgacgtt
     1861 atcccgaatc tggtcgacgc cagggacaat caaataagca gtgctgctcg tgtgtcatca
     1921 tgggtaccag tggttccgga ggatctggag gatctggtat ggtgagcaag ggcgaggagc
     1981 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
     2041 tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca
     2101 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg
     2161 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     2221 ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca
     2281 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     2341 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca
     2401 gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga
     2461 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     2521 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccaagc
     2581 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     2641 ccgggatcac tctcggcatg gacgagctgt acaagtagca tatggcggcc gcggaagggc
     2701 gaattcgacc cagctttctt gtacaaagtg gtgatatcga attcctgcag cccgggggat
     2761 ccactagttc tagagcggcc gccaccgcgg ccgctgtcat cagatcgcca tctcgcgccc
     2821 gtgcctctga cttctaagtc caattactct tcaacatccc tacatgctct ttctccctgt
     2881 gctcccaccc cctatttttg ttattatcaa aaaacttctc ttaatttctt tgttttttag
     2941 cttcttttaa gtcacctcta acaatgaaat tgtgtagatt caaaaataga attaattcgt
     3001 aataaaaagt cgaaaaaaat tgtgctccct ccccccatta ataataattc tatcccaaaa
     3061 tctacacaat gttctgtgta cacttcttat gttttttact tctgataaat ttttttgaaa
     3121 catcatagaa aaaaccgcac acaaaatacc ttatcatatg ttacgtttca gtttatgacc
     3181 gcaattttta tttcttcgca cgtctgggcc tctcatgacg tcaaatcatg ctcatcgtga
     3241 aaaagttttg gagtattttt ggaatttttc aatcaagtga aagtttatga aattaatttt
     3301 cctgcttttg ctttttgggg tttcccctat tgtttgtcaa gatttcgagg acggcgtttt
     3361 tcttgctaaa atcacaagta ttgatgagca cgatgcaaga aagatcggaa gaaggtttgg
     3421 gtttgaggct cagtggaagg tgagtagaag ttgataattt gaaagtggag tagtgtctat
     3481 ggggtttttg ccgagctcca gcttttgttc cctttagtga gggttaattg cgcgcttggc
     3541 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     3601 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     3661 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     3721 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     3781 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     3841 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     3901 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     3961 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     4021 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     4081 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     4141 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     4201 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     4261 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     4321 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     4381 ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     4441 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     4501 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     4561 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     4621 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     4681 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     4741 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     4801 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     4861 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     4921 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     4981 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     5041 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     5101 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     5161 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     5221 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     5281 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     5341 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     5401 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     5461 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     5521 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     5581 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     5641 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccac
//
