LOCUS       sequence-256716-        6852 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Dox-induced expression of SOX17.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6852)
  TITLE     Lian Lab CRISPR plasmids
REFERENCE   2  (bases 1 to 6852)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pYJ42-XLone-SOX17
FEATURES             Location/Qualifiers
     source          1..6852
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        9..376
                     /label=TRE3GS promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G, modified to eliminate
                     binding sites for endogenous mammalian transcription 
                     factors"
     protein_bind    17..35
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    53..71
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    89..107
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    125..143
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    161..179
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    197..215
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    233..251
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             1337..1354
                     /codon_start=1
                     /product="HQHQHQ metal affinity tag"
                     /label=HQ tag
                     /note="The HQ tag binds to nickel and other metal ions much
                     like the 6xHis tag, but may allow elution at lower 
                     concentrations of imidazole (Godat et al., 2008)."
                     /translation="HQHQHQ"
     CDS             1343..1360
                     /codon_start=1
                     /product="HQHQHQ metal affinity tag"
                     /label=HQ tag
                     /note="The HQ tag binds to nickel and other metal ions much
                     like the 6xHis tag, but may allow elution at lower 
                     concentrations of imidazole (Godat et al., 2008)."
                     /translation="HQHQHQ"
     polyA_signal    1814..1948
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     repeat_region   2224..2258
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(2269..2286)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2269..2285)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2278..2300)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2494..2513)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2613..2635
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2673..2691)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2759..2863
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2864..2932,2933..3724)
                     /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(3082..3101)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3895..4483
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4384..4403
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4637..4654
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4771..4792
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4807..4812,4813..4830,4831..4837)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4845..4861
                     /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     4850..4872
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4869..4885
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4869..4885
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   4911..4973
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     polyA_signal    5149..5230
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(5347..5745)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     CDS             complement(5746..5802)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             complement(5812..6555)
                     /codon_start=1
                     /product="modified rtTA protein that binds tightly to 
                     promoters containing the tet operator in the presence of 
                     doxycycline"
                     /label=Tet-On(R) 3G
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     CDS             complement(6556..6579)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      6579..6588
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(6608..6819)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
ORIGIN
        1 atcacctcga gtttactccc tatcagtgat agagaacgta tgaagagttt actccctatc
       61 agtgatagag aacgtatgca gactttactc cctatcagtg atagagaacg tataaggagt
      121 ttactcccta tcagtgatag agaacgtatg accagtttac tccctatcag tgatagagaa
      181 cgtatctaca gtttactccc tatcagtgat agagaacgta tatccagttt actccctatc
      241 agtgatagag aacgtataag ctttgcttat gtaaaccagg gcgcctataa aagagtgctg
      301 attttttgag taaacttcaa ttccacaaca cttttgtctt ataccaactt tccgtaccac
      361 ttcctaccct cgtaaaggta ccgccaccat gagcagcccg gatgcgggat acgccagtga
      421 cgaccagagc cagacccaga gcgcgctgcc cgcggtgatg gccgggctgg gcccctgccc
      481 ctgggccgag tcgctgagcc ccatcgggga catgaaggtg aagggcgagg cgccggcgaa
      541 cagcggagca ccggccgggg ccgcgggccg agccaagggc gagtcccgta tccggcggcc
      601 gatgaacgct ttcatggtgt gggctaagga cgagcgcaag cggctggcgc agcagaatcc
      661 agacctgcac aacgccgagt tgagcaagat gctgggcaag tcgtggaagg cgctgacgct
      721 ggcggagaag cggcccttcg tggaggaggc agagcggctg cgcgtgcagc acatgcagga
      781 ccaccccaac tacaagtacc ggccgcggcg gcgcaagcag gtgaagcggc tgaagcgggt
      841 ggagggcggc ttcctgcacg gcctggctga gccgcaggcg gccgcgctgg gccccgaggg
      901 cggccgcgtg gccatggacg gcctgggcct ccagttcccc gagcagggct tccccgccgg
      961 cccgccgctg ctgcctccgc acatgggcgg ccactaccgc gactgccaga gtctgggcgc
     1021 gcctccgctc gacggctacc cgttgcccac gcccgacacg tccccgctgg acggcgtgga
     1081 ccccgacccg gctttcttcg ccgccccgat gcccggggac tgcccggcgg ccggcaccta
     1141 cagctacgcg caggtctcgg actacgctgg ccccccggag cctcccgccg gtcccatgca
     1201 cccccgactc ggcccagagc ccgcgggtcc ctcgattccg ggcctcctgg cgccacccag
     1261 cgcccttcac gtgtactacg gcgcgatggg ctcgcccggg gcgggcggcg ggcgcggctt
     1321 ccagatgcag ccgcaacacc agcaccagca ccagcaccag caccaccccc cgggccccgg
     1381 acagccgtcg ccccctccgg aggcactgcc ctgccgggac ggcacggacc ccagtcagcc
     1441 cgccgagctc ctcggggagg tggaccgcac ggaatttgaa cagtatctgc acttcgtgtg
     1501 caagcctgag atgggcctcc cctaccaggg gcatgactcc ggtgtgaatc tccccgacag
     1561 ccacggggcc atttcctcgg tggtgtccga cgccagctcc gcggtatatt actgcaacta
     1621 tcctgacgtg tagactagta gaccacctcc cctgcgagct aagctggaca gccaatgacg
     1681 ggtaagagag tgacattttt cactaaccta agacaggagg gccgtcagag ctactgccta
     1741 atccaaagac gggtaaaagt gataaaaatg tatcactcca acctaagaca ggcgcagctt
     1801 ccgagggatt tgagatccag acatgataag atacattgat gagtttggac aaaccaaaac
     1861 tagaatgcag tgaaaaaaat gccttatttg tgaaatttgt gatgctattg ccttatttgt
     1921 aaccattata agctgcaata aacaagtttg atatctataa caagaaaata tatatataat
     1981 aagttatcac gtaagtagaa catgaaataa caatataatt atcgtatgag ttaaatctta
     2041 aaagtcacgt aaaagataat catgcgtcat tttgactcac gcggtcgtta tagttcaaaa
     2101 tcagtgacac ttaccgcatt gacaagcacg cctcacggga gctccaagcg gcgactgaga
     2161 tgtcctaaat gcacagcgac ggattcgcgc tatttagaaa gagagagcaa tatttcaaga
     2221 atgcatgcgt caattttacg cagactatct ttctagggtt aagaattcac tggccgtcgt
     2281 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     2341 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     2401 gttgcgcagc ctgaatggcg aatggcgcct gatgcggtat tttctcctta cgcatctgtg
     2461 cggtatttca caccgcatat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
     2521 aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc
     2581 ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc
     2641 accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt
     2701 taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg
     2761 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     2821 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     2881 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     2941 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     3001 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     3061 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     3121 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     3181 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     3241 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     3301 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     3361 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     3421 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     3481 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     3541 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     3601 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     3661 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     3721 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     3781 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     3841 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     3901 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     3961 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     4021 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     4081 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     4141 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     4201 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     4261 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     4321 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     4381 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     4441 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     4501 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     4561 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     4621 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
     4681 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
     4741 ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc
     4801 ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca
     4861 atttcacaca ggaaacagct atgaccatga ttacgccaag gtcgacttaa ccctagaaag
     4921 ataatcatat tgtgacgtac gttaaagata atcatgcgta aaattgacgc atgtgtttta
     4981 tcggtctgta tatcgaggtt tatttattaa tttgaataga tattaagttt tattatattt
     5041 acacttacat actaataata aattcaacaa acaatttatt tatgtttatt tatttattaa
     5101 aaaaaaacaa aaactcaaaa tttcttctat aaagtaacaa aacttttagc agtgaaaaaa
     5161 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     5221 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg
     5281 ggaggttttt taaagcaagt aaaacctcta caaatgtggt atggctgatt atgatcctct
     5341 ggagatttag ccctcccaca cataaccaga gggcagcaat tcacgaatcc caactgccgt
     5401 cggctgtcca tcactgtcct tcactatggc tttgatccca ggatgcagat cgagaagcac
     5461 ctgtcggcac cgtccgcagg ggctcaagat gcccctgttc tcatttccga tcgcgacgat
     5521 acaagtcagg ttgccagctg ccgcagcagc agcagtgccc agcaccacga gttctgcaca
     5581 aggtccccca gtaaaatgat atacattgac accagtgaag atgcggccgt cgctagagag
     5641 agctgcgctg gcgacgctgt agtcttcaga gatggggatg ctgttgattg tagccgttgc
     5701 tctttcaatg agggtggatt cttcttgaga caaaggcttg gccatggggc cggggttctc
     5761 ctccacgtcg ccggcctgct tcagcaggct gaagttggtg gcgccgctgc ccccggggag
     5821 catgtcaagg tcaaaatcgt caagagcgtc agcaggcagc atatcaaggt caaagtcgtc
     5881 aagggcatcg gctgggagca tgtctaagtc aaaatcgtca agggcgtcgg tcggcccgcc
     5941 gctttcgcac tttagctgtt tctccaggcc acatatgatt agttccaggc cgaaaaggaa
     6001 ggcaggttcg gctccctgcc ggtcgaacag ctcaattgct tgtttcagaa gtgggggcat
     6061 agaatcggtg gtaggtgtct ctctttcctc ttttgctact tgatgctcct gttcctccaa
     6121 tacgcagccc agtgtaaagt ggcccacggc ggacagagcg tacagtgcgt tctccaggga
     6181 gaagccttgc tgacacagga acgcgagctg attttccagg gtttcgtact gtttctctgt
     6241 tgggcgggtg ccgagatgca ctttagcccc gtcgcgatgt gagaggagag cacagcggta
     6301 tgacttggcg ttgttccgca gaaagtcttg ccatgactcg ccttccaggg ggcaggagtg
     6361 ggtatgatgc ctgtccagca tctcgattgg cagggcatcg agcagggccc gcttgttctt
     6421 cacgtgccag tacagggtag gctgctcaac tcccagcttt tgagcgagtt tccttgtcgt
     6481 caggccttcg ataccgactc cattgagtaa ttccagagca gagtttatga ctttgctctt
     6541 gtccagtcta gacatcttat cgtcatcgtc tttgtaatcc atggtggcgg atcccgcgtc
     6601 acgacacctg tgttctggcg gcaaacccgt tgcgaaaaag aacgttcacg gcgactactg
     6661 cacttatata cggttctccc ccaccctcgg gaaaaaggcg gagccagtac acgacatcac
     6721 tttcccagtt taccccgcgc caccttctct aggcaccggt tcaattgccg acccctcccc
     6781 ccaacttctc ggggactgtg ggcgatgtgc gctctgccca ctgacgggca ccggagccac
     6841 tcgagtggaa tt
//
