LOCUS       sequence-325306-        8076 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses Sonic Hedgehog.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8076)
  AUTHORS   Desai PB, Stuck MW, Lv B, Pazour GJ
  TITLE     Ubiquitin links smoothened to intraflagellar transport to regulate 
            Hedgehog signaling.
  JOURNAL   J Cell Biol. 2020 Jul 6;219(7). pii: 151798. doi: 
            10.1083/jcb.201912104.
  PUBMED    32435793
REFERENCE   2  (bases 1 to 8076)
  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: "J Cell 
            Biol."; date: "2020-07-6"; volume: "219(7). pii: 151798. doi"; 
            pages: " 10.1083/jcb.201912104"
COMMENT     Sequence Label: MS03
FEATURES             Location/Qualifiers
     source          1..8076
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..634
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    681..806
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1303..1536
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1721..1765
                     /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    2061..2178
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        2230..2533
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2534..2737
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2688..2708
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2734..2758
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     misc_feature    join(3460..4033,4034..4036,4037..4045)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3626..3643)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3853..3872
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4046..4441
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     CDS             4451..4504
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     misc_feature    4518..5106
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4571..4591)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4989..5000)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             5254..5434
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(5481..5499)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5567..5671
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5672..5740,5741..6532)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5890..5909)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6703..7291
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7445..7462
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7702..7723
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7738..7743,7744..7761,7762..7768)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7776..7792
                     /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     7781..7803
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7800..7816
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7800..7816
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7866..7885
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca
       61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac
      121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca
      181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg
      241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag
      301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg
      361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat
      421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga
      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
      541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag
      661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg
      721 caagaggcga ggggcggcga ctgcagagta cgccaaaaat tttgactagc ggaggctaga
      781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg
      841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg
      901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct
      961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat
     1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca
     1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc
     1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag
     1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccgaa ttctcaacaa
     2041 taggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg
     2101 gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt
     2161 acaaaaattc aaaattttcg ggtttattac tcggacagca gagatccagt ttggactagt
     2221 ggagttccgc gttacataac ttacgcaaaa tggcccgcct ggctgaccgc ccaacgaccc
     2281 ccgcccattg acgtcaataa tgaccaatgt tcccatagta acgccaatag ggactttcca
     2341 ttgacgtcaa tgggtggagt atttacgcaa aactgcccac ttggcagtac atcaagtgta
     2401 tcatttgcca acaacgcccc ctattgacgt caatgacgca aaatggcccg cctggcatta
     2461 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     2521 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
     2581 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
     2641 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
     2701 taggcgtgta cggtgggagg tctatataag cagagctcgt ttagtgaacc gtcagatcgc
     2761 ctggagacgc catccacgct gttttgacct ccattcaaga caccggcggc cttaattaat
     2821 gatcaggatc caccatgctg ctgctggcga gatgtctgct gctagtcctc gtctcctcgc
     2881 tgctggtatg ctcgggactg gcgtgcggac cgggcagggg gttcgggaag aggaggcacc
     2941 ccaaaaagct gaccccttta gcctacaagc agtttatccc caatgtggcc gagaagaccc
     3001 taggcgccag cggaaggtat gaagggaaga tctccagaaa ctccgagcga tttaaggaac
     3061 tcacccccaa ttacaacccc gacatcatat ttaaggatga agaaaacacc ggagcggaca
     3121 ggctgatgac tcagaggtgt aaggacaagt tgaacgcttt ggccatctcg gtgatgaacc
     3181 agtggccagg agtgaaactg cgggtgaccg agggctggga cgaagatggc caccactcag
     3241 aggagtctct gcactacgag ggccgcgcag tggacatcac cacgtctgac cgcgaccgca
     3301 gcaagtacgg catgctggcc cgcctggcgg tggaggccgg cttcgactgg gtgtactacg
     3361 agtccaaggc acatatccac tgctcggtga aagcagagaa ctcggtggcg gccaaatcgg
     3421 gaggctaatc tagagcggcc gctaagtaag taagatccgc ccctctccct ccccccccct
     3481 aacgttactg gccgaagccg cttggaataa ggccggtgtg cgtttgtcta tatgttattt
     3541 tccaccatat tgccgtcttt tggcaatgtg agggcccgga aacctggccc tgtcttcttg
     3601 acgagcattc ctaggggtct ttcccctctc gccaaaggaa tgcaaggtct gttgaatgtc
     3661 gtgaaggaag cagttcctct ggaagcttct tgaagacaaa caacgtctgt agcgaccctt
     3721 tgcaggcagc ggaacccccc acctggcgac aggtgcctct gcggccaaaa gccacgtgta
     3781 taagatacac ctgcaaaggc ggcacaaccc cagtgccacg ttgtgagttg gatagttgtg
     3841 gaaagagtca aatggctctc ctcaagcgta ttcaacaagg ggctgaagga tgcccagaag
     3901 gtaccccatt gtatgggatc tgatctgggg cctcggtaca catgctttac atgtgtttag
     3961 tcgaggttaa aaaaacgtct aggccccccg aaccacgggg acgtggtttt cctttgaaaa
     4021 acacgatgat aatatggcca caaccatggc caagcctttg tctcaagaag aatccaccct
     4081 cattgaaaga gcaacggcta caatcaacag catccccatc tctgaagact acagcgtcgc
     4141 cagcgcagct ctctctagcg acggccgcat cttcactggt gtcaatgtat atcattttac
     4201 tgggggacct tgtgcagaac tcgtggtgct gggcactgct gctgctgcgg cagctggcaa
     4261 cctgacttgt atcgtcgcga tcggaaatga gaacaggggc atcttgagcc cctgcggacg
     4321 gtgccgacag gtgcttctcg atctgcatcc tgggatcaaa gccatagtga aggacagtga
     4381 tggacagccg acggcagttg ggattcgtga attgctgccc tctggttatg tgtgggaggg
     4441 cggtagtggc gagggcagag gaagtcttct aacatgcggt gacgtggagg agaatcccgg
     4501 ccctctcgag agatcctaat caacctctgg attacaaaat ttgtgaaaga ttgactggta
     4561 ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc
     4621 atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt
     4681 ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg
     4741 ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt
     4801 tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct
     4861 ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg aaatcatcgt
     4921 cctttccttg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct
     4981 acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc
     5041 ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct
     5101 ccccgcctgg tacctttaag accaatgact tacaaggcag ctgtagatct tagccacttt
     5161 ttaaaagaaa aggggggact ggaagggcta attcactccc aacgaagaca agatcacctg
     5221 caggacaggc gcgccctgct ttttgcttgt actgggtctc tctggttaga ccagatctga
     5281 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
     5341 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
     5401 agaccctttt agtcagtgtg gaaaatctct agcacccggg cgattaagga aagggctaga
     5461 tcattcttga agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     5521 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     5581 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     5641 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     5701 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     5761 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     5821 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     5881 taaagttctg ctatgtggcg cggtattatc ccgtgttgac gccgggcaag agcaactcgg
     5941 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     6001 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     6061 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     6121 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     6181 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     6241 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     6301 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     6361 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     6421 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     6481 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     6541 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     6601 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     6661 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     6721 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     6781 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     6841 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     6901 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     6961 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     7021 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     7081 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     7141 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     7201 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     7261 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     7321 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     7381 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     7441 gcgcagcgag tcagtgagcg aggaagcgga agagcgccca atacgcaaac cgcctctccc
     7501 cgcgcgttgg ccgattcatt aatgcagcaa gctcatggct gactaatttt ttttatttat
     7561 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     7621 ggaggcctag gcttttgcaa aaagctcccc gtggcacgac aggtttcccg actggaaagc
     7681 gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt
     7741 acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac
     7801 aggaaacagc tatgacatga ttacgaattt cacaaataaa gcattttttt cactgcattc
     7861 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggatca actggataac
     7921 tcaagctaac caaaatcatc ccaaacttcc caccccatac cctattacca ctgccaatta
     7981 cctgtggttt catttactct aaacctgtga ttcctctgaa ttattttcat tttaaagaaa
     8041 ttgtatttgt taaatatgta ctacaaactt agtagt
//
