LOCUS       sequence-211334-        7209 bp    DNA     circular SYN 14-APR-2025
DEFINITION  reporter of Hedgehog signaling activity.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7209)
  AUTHORS   Li T, Fan J, Blanco-Sanchez B, Giagtzoglou N, Lin G, Yamamoto S, 
            Jaiswal M, Chen K, Zhang J, Wei W, Lewis MT, Groves AK, Westerfield 
            M, Jia J, Bellen HJ
  TITLE     Ubr3, a Novel Modulator of Hh Signaling Affects the Degradation of 
            Costal-2 and Kif7 through Poly-ubiquitination.
  JOURNAL   PLoS Genet. 2016 May 19;12(5):e1006054. doi: 
            10.1371/journal.pgen.1006054. eCollection 2016 May.
  PUBMED    27195754
REFERENCE   2  (bases 1 to 7209)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "PLoS 
            Genet."; date: "2016-05-19"; volume: "12(5):e1006054. doi"; pages: "
            10.1371/journal.pgen.1006054. eCollection 2016 May"
COMMENT     Sequence Label: 7Gli:GFP
FEATURES             Location/Qualifiers
     source          1..7209
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      414..423
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(420..422,423..425,426..1139)
                     /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(420..422,423..425,426..1136)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(465..486)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(726..745)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1073..1094
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1164..1752
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1217..1237)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1635..1646)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             1839..2072
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2144..2265
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2181..2200)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2235..2254
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2305..2440
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2460..2479)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2461..2479)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2489..2506)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2489..2505)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2498..2520)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2647..3102
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2734..2753)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2944..2965
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3128..3232
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3233..3301,3302..4093)
                     /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(3451..3470)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4264..4852
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4753..4772
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5006..5023
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5140..5161
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5176..5181,5182..5199,5200..5206)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5214..5230
                     /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     5219..5241
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5238..5254
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5238..5254
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5273..5293
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5275..5293
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5321..5547
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5548..5728
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5775..5900
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6393..6626
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6811..6855
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    join(7122..7209,1..30)
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 caaaaacaaa ttacaaaaat tcaaaatttt atcgatcacg agactagcct cctagcccgg
       61 gctcgagcag ctgaagcttg catgccctgc agcgacctcg acaagcaggg aacacccaag
      121 tagaagttcc gacaagcagg gaacacccaa gtagaagctc tcgacctcga caagcaggga
      181 acacccaagt agaagctccg acaagcaggg aacacccaag tagaagctct cgacctcgac
      241 aagcagggaa cacccaagta gaagctccga caagcaggga acacccaagt agaagctctc
      301 gacctcgaca agcagggaac acccaagtag aagctctgca gagggtatat aatggatccg
      361 gtactcgaga tctgcgatct aagtaagctt ggcattccgg tactgttggt aaagccacca
      421 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
      481 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
      541 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
      601 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc
      661 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
      721 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
      781 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
      841 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
      901 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
      961 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     1021 acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     1081 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtaaa
     1141 gcggccgcga ctctagagtc gacaatcaac ctctggatta caaaatttgt gaaagattga
     1201 ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt
     1261 tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt
     1321 tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg
     1381 tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg
     1441 ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc
     1501 gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaagc
     1561 tgacgtcctt tccatggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct
     1621 tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg
     1681 ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg
     1741 ccgcctcccc gcctggaatt cgagctcggt acctttaaga ccaatgactt acaaggcagc
     1801 tgtagatctt agccactttt taaaagaaaa ggggggactg gaagggctaa ttcactccca
     1861 acgaagacaa gatctgcttt ttgcttgtac tgggtctctc tggttagacc agatctgagc
     1921 ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg
     1981 agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag
     2041 acccttttag tcagtgtgga aaatctctag cagtagtagt tcatgtcatc ttattattca
     2101 gtatttataa cttgcaaaga aatgaatatc agagagtgag aggaacttgt ttattgcagc
     2161 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     2221 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctggctcta
     2281 gctatcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     2341 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct
     2401 gagctattcc agaagtagtg aggaggcttt tttggaggcc tagggacgta cccaattcgc
     2461 cctatagtga gtcgtattac gcgcgctcac tggccgtcgt tttacaacgt cgtgactggg
     2521 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
     2581 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
     2641 aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     2701 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     2761 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     2821 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     2881 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     2941 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     3001 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     3061 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg
     3121 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     3181 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     3241 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     3301 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     3361 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     3421 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     3481 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     3541 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     3601 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     3661 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     3721 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     3781 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     3841 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     3901 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     3961 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     4021 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     4081 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     4141 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     4201 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     4261 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     4321 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     4381 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     4441 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     4501 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     4561 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     4621 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     4681 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     4741 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     4801 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     4861 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     4921 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     4981 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc
     5041 aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag
     5101 gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca
     5161 ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag
     5221 cggataacaa tttcacacag gaaacagcta tgaccatgat tacgccaagc gcgcaattaa
     5281 ccctcactaa agggaacaaa agctggagct gcaagcttaa tgtagtctta tgcaatactc
     5341 ttgtagtctt gcaacatggt aacgatgagt tagcaacatg ccttacaagg agagaaaaag
     5401 caccgtgcat gccgattggt ggaagtaagg tggtacgatc gtgccttatt aggaaggcaa
     5461 cagacgggtc tgacatggat tggacgaacc actgaattgc cgcattgcag agatattgta
     5521 tttaagtgcc tagctcgata cataaacggg tctctctggt tagaccagat ctgagcctgg
     5581 gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg
     5641 cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc
     5701 ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa cagggacttg aaagcgaaag
     5761 ggaaaccaga ggagctctct cgacgcagga ctcggcttgc tgaagcgcgc acggcaagag
     5821 gcgaggggcg gcgactggtg agtacgccaa aaattttgac tagcggaggc tagaaggaga
     5881 gagatgggtg cgagagcgtc agtattaagc gggggagaat tagatcgcga tgggaaaaaa
     5941 ttcggttaag gccaggggga aagaaaaaat ataaattaaa acatatagta tgggcaagca
     6001 gggagctaga acgattcgca gttaatcctg gcctgttaga aacatcagaa ggctgtagac
     6061 aaatactggg acagctacaa ccatcccttc agacaggatc agaagaactt agatcattat
     6121 ataatacagt agcaaccctc tattgtgtgc atcaaaggat agagataaaa gacaccaagg
     6181 aagctttaga caagatagag gaagagcaaa acaaaagtaa gaccaccgca cagcaagcgg
     6241 ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag tgaattatat
     6301 aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc aaagagaaga
     6361 gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg gttcttggga
     6421 gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc cagacaatta
     6481 ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc gcaacagcat
     6541 ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct ggctgtggaa
     6601 agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa actcatttgc
     6661 accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca gatttggaat
     6721 cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt aatacactcc
     6781 ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt ggaattagat
     6841 aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta tataaaatta
     6901 ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt actttctata
     6961 gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct cccaaccccg
     7021 aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga cagagacaga
     7081 tccattcgat tagtgaacgg atctcgacgg tatcggttaa cttttaaaag aaaagggggg
     7141 attggggggt acagtgcagg ggaaagaata gtagacataa tagcaacaga catacaaact
     7201 aaagaatta
//
