LOCUS       sequence-258787-        8666 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Prgef-1 INS-4::GFP unc-54 3' UTR C.elegans pan-neural expression of 
            INS-4::GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8666)
  AUTHORS   Hung WL, Wang Y, Chitturi J, Zhen M
  TITLE     A Caenorhabditis elegans developmental decision requires insulin 
            signaling-mediated neuron-intestine communication.
  JOURNAL   Development. 2014 Apr;141(8):1767-79. doi: 10.1242/dev.103846. Epub 
            2014 Mar 26.
  PUBMED    24671950
REFERENCE   2  (bases 1 to 8666)
  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"; doi: 
            "10.1242/dev.103846"; journalName: "Development"; date: "2014-04"; 
            volume: "141"; issue: "8"; pages: "1767-79"
COMMENT     Sequence Label: pJH2255
FEATURES             Location/Qualifiers
     source          1..8666
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    78..99
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(114..119,120..137,138..144)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    152..168
                     /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)."
     primer_bind     157..179
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     176..192
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     176..192
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     regulatory      4099..4108
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4105..4107,4108..4110,4111..4824)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(4105..4107,4108..4110,4111..4821)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4150..4171)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4411..4430)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4758..4779
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(4849..4873)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             4899..4940
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(5766..5783)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5766..5782)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5775..5797)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5995..6450
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6082..6101)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6292..6313
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6467..6486)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6586..6608
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6646..6664)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6732..6836
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6837..6905,6906..7697)
                     /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     complement(7055..7074)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7868..8456
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8357..8376
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8610..8627
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc
       61 agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac
      121 tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga
      181 aacagctatg acatgattac gaattcgaat ggccatggga cgtcgacgct tgcatgcctg
      241 cagcgtttcc gataccccct tatatcagca cacattcagt catcatacca caaaatgttg
      301 tgatagtgat aggcaaaact accgtatttc ctctattaat cttgcatgca agactaattt
      361 tcgattaacc cgtaggggtg caagactaat agagactgca agactattag aggctgaaat
      421 actaattttc gtatgctcaa taattttgga aattggccta ttttttgtag aaacttgata
      481 ccgtttaaac aaggaaaaat acacactttt taatatttca tcaataattt gaacgatttt
      541 gtgattttaa gttcaatttg cccaaaaaaa gacaattttt ctgacgttac cgcaaagtaa
      601 tgctggtcag gcaaaaagag cggtgcaaaa atataagaga ctgcaatact aatagaggaa
      661 atacggtaat tgaattttag tgaaacttgc gacagttttc ctcatttttt tgttattccg
      721 gcatacgagt gtggcatacg agtgaggtca tcttttgttt cttccgtttc ttcatcgctt
      781 ttgaaaaaaa atgttgaaga aaacttcgta gccgatagcc ggataatgtt tggcacggcg
      841 ttccaacata tatcattgga aattttttaa gatttttgcg aaaaatcaca ttcttcacga
      901 tgagaacacg ttattgaagg aatataaatc aagaataaca tatagttata tttctctatt
      961 actttaacgt taaatatgag caaatttgag cattttgatt gcgatgaaaa gcagaatcga
     1021 gtcaactgaa atccgttcaa aaacctaagc tcggttcgcc cgaaagtcga tttttccaat
     1081 agccgaacag cactgcttct ttattttcca ggtatttttg gggtgtaaga attatatctt
     1141 ctttgtttgg gtatttacgt ttccgaattc cctttcttga tctgtcgcta tttttcagaa
     1201 accagaacag tttatccttt ttcatgagaa cctgaatagc tcaaaaaccc gtttaatttc
     1261 ttttcatcat ctcatttgaa actttcctag tcttcttaat gatttccagg ctcctcctac
     1321 atttgtatct caagatgcat acactttatt tccgctccaa ttcgaatcat ctacttctct
     1381 tttttcttca actttctact tttttccatt cttttctttc aaatgttcga cgtcttcaag
     1441 tgactgctct tcattctcct catttttctc gaatttcatt cttgtctctt ttttgctcaa
     1501 aaatggaaaa tgaaagtgac gtgagatttg gacggcggga cacgggggca gtagaagcag
     1561 caaaaaggag agaaagagga cacaaataag aagaacgaat tcaaaaataa gcggagagga
     1621 gctatttccg tcaattctac ctccccaatc ttcatcaatt cggctcaatt gaatgacgtc
     1681 acagggagat aaaccgtttg gatgagcgcc gtccatcact gacgccatcc cgtttgggac
     1741 aagaaaagag aaaaaagagc acaaagtttt tggtgacgga tcttgtcaat catatgaaag
     1801 ttgttctgat tgattgtcag tttttttcct acttttttgg attctatcca cttctgaact
     1861 tttgacaagt ttcaaacttt ctgaatcatt ttctatgcat tttcctggaa ttctttttat
     1921 gtaaaatatg aaatagaatg tttttgaatt caagttctgc ttttttcttc ttttctgttc
     1981 tgttcgggct tgggtatgct ttttttaaaa attattttgc acatcgacca ataagtgcgc
     2041 aacttataaa attaatttat ttttgttaat ttttgaaata cttgtattgc tttaagtgat
     2101 ctgacctcgc cctgagcttt ccacgtagtt atcaaataca aatcctccaa gggtaacgta
     2161 cctatattac tgatctttat aataacttta tcacctgtcc agttccagag gaattctgtt
     2221 aagcttatag gcacagaagg agtcatttct gctggttttt aatgatccaa atctttattt
     2281 caagtaaaaa actgaacact tgcgaataaa actatcagat taaccattca ccaaaaatgt
     2341 gtttgaatct aaaactttct cagtattcca aatatagaaa taaataacca cgacattgct
     2401 aaaatctgtc tgaattgtgt actccttacc gtgaagtaat aatggatata atgaatcgtt
     2461 tgaaatgaat gatcagcaca tttttggtga aagatcacaa ataaggaata agcgtcggaa
     2521 aataaaacga ttatttcgga tcaaaaattt gttgaagatc atatacactc gagaccaaga
     2581 ttattctaga caattttcaa attggcttct ttgtttgcaa atcttcataa taatctgtga
     2641 agtttggaaa tttgaatttt taatcttttt tcatagatta tagtttttat ttctttgcaa
     2701 aactatatta aaaaccgatg cattgtttta gggaaattaa tgagcctttt gttcaacact
     2761 aaaaacaata aaattaaaat tttggcttca tcatttgacc ttttttaagt tcgaaaactt
     2821 ttctcgtatt tctgaaccgc caattttttc acacatctct agacttttgg tgcccgttcc
     2881 agaaagttaa gtaattgcta ttctaagaaa gttctcaaca ttgtttttta gttctgattg
     2941 aattctgatg ttccaggaat atattttaaa tttaatattc tttgcactat ttctatacta
     3001 actaaataat aaatagtcta agtatgttca atgagaacaa aaaagctcta tctatatttt
     3061 tctatcctat ttcatttata tccttttcat tttgaactca ccagcttctc ttcttcttct
     3121 tgtccatata cttcttaaaa gtctcgaact tctctctctc actctccatc aatttcttta
     3181 tggctaccgc ttgcgctctg ccgcttccga agaaagaagg cgttggcaga gctctcaatt
     3241 ctattttttc tcgccgtagg ctatcgattt ctcagcttct cttctctctt cctcactcca
     3301 ccaccaccac ttcgggcttc ttcttttctt cattttcgtc ttcttcttca tcctcttctt
     3361 tttttttcag ttccccctca ctcctcccct tatatgcgcg tgcgagaggg gtgcaaaagc
     3421 agcgcgcatc cgagaattga aacgagaaaa cggagacgca gcagcagttc gtcctcagaa
     3481 aaatagccag taaaaagaga aaaagataga gagagaccta tcgcatttta ttttcaaatt
     3541 gcaattcctg caattcatgt gtgcgtgtgc ccattttcaa ttcttcccgt tatttttcag
     3601 accattacca acgttctttc tagatcttga gacaatcttc cttctgctca atcgttcgtc
     3661 gtgaagacgg catcgacgac gacgacgatc agcagtaaag gatccccatg ttttcattct
     3721 ttacatattt ccttctctcc gcacttcttc tctccgcttc atgtcgacaa ccttccatgg
     3781 acaccagcaa agccgatcgt attctacgag agatcgaaat ggaaacagaa ctcgaaaatc
     3841 aactctcccg agcacgacga gtcccagctg gagaggttcg tgcctgtgga agacgacttc
     3901 ttctctttgt ctggtcaacc tgtggagaac catgcacgcc acgtacgttt aatgctaaaa
     3961 attttttaaa aaaccttaat ctgaatttat tatttttcag aagaggacat ggacattgcc
     4021 acagtttgct gcacaacaca gtgcactcca tcatatataa aacaagcttg ctgcccagaa
     4081 aagggggatc caccggtcgc caccatggtg agcaagggcg aggagctgtt caccggggtg
     4141 gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc
     4201 gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc
     4261 aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc
     4321 agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc
     4381 tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag
     4441 gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag
     4501 gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat
     4561 atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc
     4621 gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc
     4681 cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc
     4741 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc
     4801 ggcatggacg agctgtacaa gtaaagcggc cgcactcgag atatctagac ccagctttct
     4861 tgtacaaagt ggttgatcta gagggcccgc ggttcgaagg taagcctatc cctaaccctc
     4921 tcctcggtct cgattctacg cgtaccggtt agtaatgagt ttatctgagc tccgcatcgg
     4981 ccgctgtcat cagatcgcca tctcgcgccc gtgcctctga cttctaagtc caattactct
     5041 tcaacatccc tacatgctct ttctccctgt gctcccaccc cctatttttg ttattatcaa
     5101 aaaacttctc ttaatttctt tgttttttag cttcttttaa gtcacctcta acaatgaaat
     5161 tgtgtagatt caaaaataga attaattcgt aataaaaagt cgaaaaaaat tgtgctccct
     5221 ccccccatta ataataattc tatcccaaaa tctacacaat gttctgtgta cacttcttat
     5281 gttttttact tctgataaat ttttttgaaa catcatagaa aaaaccgcac acaaaatacc
     5341 ttatcatatg ttacgtttca gtttatgacc gcaattttta tttcttcgca cgtctgggcc
     5401 tctcatgacg tcaaatcatg ctcatcgtga aaaagttttg gagtattttt ggaatttttc
     5461 aatcaagtga aagtttatga aattaatttt cctgcttttg ctttttgggg tttcccctat
     5521 tgtttgtcaa gatttcgagg acggcgtttt tcttgctaaa atcacaagta ttgatgagca
     5581 cgatgcaaga aagatcggaa gaaggtttgg gtttgaggct cagtggaagg tgagtagaag
     5641 ttgataattt gaaagtggag tagtgtctat ggggtttttg ccttaaatga cagaatacat
     5701 tcccaatata ccaaacataa ctgtttctac tagtggccac gtgggccgtg caccttaagc
     5761 ttggcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt
     5821 aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc
     5881 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat ggcgcctgat gcggtatttt
     5941 ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca tagtacgcgc
     6001 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     6061 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     6121 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     6181 tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc
     6241 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     6301 tgttccaaac tggaacaaca ctcaacccta tctcgggcta ttcttttgat ttataaggga
     6361 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     6421 attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca atctgctctg
     6481 atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
     6541 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
     6601 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc
     6661 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     6721 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     6781 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     6841 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     6901 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     6961 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     7021 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     7081 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     7141 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     7201 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     7261 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     7321 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     7381 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     7441 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     7501 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     7561 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     7621 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     7681 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     7741 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     7801 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     7861 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     7921 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     7981 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     8041 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     8101 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     8161 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     8221 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     8281 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     8341 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     8401 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     8461 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     8521 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     8581 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     8641 gcccaatacg caaaccgcct ctcccc
//
