LOCUS       sequence-416608-        6842 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of N-terminally EGFP-tagged NIP45 delta-SLD1. 
            Use with Flp-In T-Rex system..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6842)
  AUTHORS   Hertz EPT, Vega IA, Kruse T, Wang Y, Hendriks IA, Bizard AH, 
            Eugui-Anta A, Hay RT, Nielsen ML, Nilsson J, Hickson ID, Mailand N
  TITLE     The SUMO-NIP45 pathway processes toxic DNA catenanes to prevent 
            mitotic failure.
  JOURNAL   Nat Struct Mol Biol. 2023 Jul 20. doi: 10.1038/s41594-023-01045-0.
  PUBMED    37474739
REFERENCE   2  (bases 1 to 6842)
  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"; journalName: "Nat Struct 
            Mol Biol. 2023 Jul 20. doi: 10.1038/s41594-023-01045-0."
COMMENT     Sequence Label: EGFP-NIP45 delta-SLD1
FEATURES             Location/Qualifiers
     source          1..6842
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(183..287)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(362..381)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        553..932
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        933..1136
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1087..1107
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    1138..1156
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1159..1177
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1187..1211
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             join(1307..1309,1310..1312,1313..2023)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1307..1309,1310..1312,1313..2023)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1352..1373)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1613..1632)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1960..1981
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3112..3129)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3118..3342
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3475..3494)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3626..3673
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             3681..4703
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3681..4703
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    4833..4954
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4870..4889)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4924..4943
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5003..5019)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5003..5019)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5016..5038)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5027..5043
                     /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(5051..5057,5058..5075,5076..5081))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5096..5117
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5234..5251)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5405..5993)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5485..5504)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6164..6842,1..113,114..182))
                     /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     6787..6806
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
       61 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
      121 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
      181 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
      241 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
      301 aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca
      361 gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg
      421 aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca
      481 attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca
      541 gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat
      601 tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg
      661 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa
      721 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact
      781 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta
      841 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt
      901 acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg
      961 ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg
     1021 ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc
     1081 cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctctcc
     1141 ctatcagtga tagagatctc cctatcagtg atagagatcg tcgacgagct cgtttagtga
     1201 accgtcagat cgcctggaga cgccatccac gctgttttga cctccataga agacaccggg
     1261 accgatccag cctccggact ctagcgttta aacttaagct tccaccatgg tgagcaaggg
     1321 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg
     1381 ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct
     1441 gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct
     1501 gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt
     1561 caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg
     1621 caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga
     1681 gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa
     1741 ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa
     1801 cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca
     1861 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca
     1921 gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt
     1981 gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagaagcttg gtaccatggc
     2041 ggagcctgtg gggaagcggg gccgctggtc cggaggtagc ggtgccggcc gagggggtcg
     2101 gggcggctgg ggcggtcggg gccggcgtcc tcgggcccag cggtctccat cccggggcac
     2161 gctggacgta gtgtctgtgg acttggtcac cgacagcgat gaggaaattc tggaggtcgc
     2221 caccgctcgc ggtgccgcgg acgaggttga ggtggagccc ccggagcccc cggggccggt
     2281 cgcgtcccgg gataacagca acagtgacag cgaaggggag gacaggcggc ccgcaggacc
     2341 cccgcgggag ccggtcaggc ggcggcggcg gctggtgctg gatccggggg aggcgccgct
     2401 ggttccggtg tactcgggga aggttaaaag cagccttcgc cttatcccag atgatctatc
     2461 cctcctgaaa ctctaccctc caggggatga ggaagaggca gagctggcag attcgagtgg
     2521 tctctaccat gagggctccc catcaccagg ctctccctgg aagacaaagc tgaggactaa
     2581 ggataaagaa gagaagaaaa agacagagtt tctggatctg gacaactctc ctctgtcccc
     2641 accttcacca aggaccaaaa gcagaacgca tactcgggca ctcaagaagt taagtgaggt
     2701 gaacaagcgc ctccaggatc tccgttcctg tctgagcccc aagccacctc agggtcaaga
     2761 gcaacagggc caagaggatg aagtggtctt ggtggaaggg cccaccctcc caacaagttc
     2821 tccagaggcc acagagacgt cccaacagct ccagctccgg gtgcagggaa aggagaaaca
     2881 ccagacactg gaagtctcac tgtctcgaga ttcccctcta aagaccctca tgtcccacta
     2941 tgaggaggcc atgggactgt cgggacggaa gctctccttc ttctttgatg ggacaaagct
     3001 ttcaggcagg gagctgccag ctgacctggg catggaatct ggggacctca ttgaggtctg
     3061 gggctgagcg gccgctcgag tctagagggc ccgtttaaac ccgctgatca gcctcgactg
     3121 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
     3181 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
     3241 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
     3301 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa
     3361 ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg
     3421 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     3481 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     3541 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     3601 attagggtga tggttcacgt acctagaagt tcctattccg aagttcctat tctctagaaa
     3661 gtataggaac ttccttggcc aaaaagcctg aactcaccgc gacgtctgtc gagaagtttc
     3721 tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct ctcggagggc gaagaatctc
     3781 gtgctttcag cttcgatgta ggagggcgtg gatatgtcct gcgggtaaat agctgcgccg
     3841 atggtttcta caaagatcgt tatgtttatc ggcactttgc atcggccgcg ctcccgattc
     3901 cggaagtgct tgacattggg gaattcagcg agagcctgac ctattgcatc tcccgccgtg
     3961 cacagggtgt cacgttgcaa gacctgcctg aaaccgaact gcccgctgtt ctgcagccgg
     4021 tcgcggaggc catggatgcg atcgctgcgg ccgatcttag ccagacgagc gggttcggcc
     4081 cattcggacc gcaaggaatc ggtcaataca ctacatggcg tgatttcata tgcgcgattg
     4141 ctgatcccca tgtgtatcac tggcaaactg tgatggacga caccgtcagt gcgtccgtcg
     4201 cgcaggctct cgatgagctg atgctttggg ccgaggactg ccccgaagtc cggcacctcg
     4261 tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa tggccgcata acagcggtca
     4321 ttgactggag cgaggcgatg ttcggggatt cccaatacga ggtcgccaac atcttcttct
     4381 ggaggccgtg gttggcttgt atggagcagc agacgcgcta cttcgagcgg aggcatccgg
     4441 agcttgcagg atcgccgcgg ctccgggcgt atatgctccg cattggtctt gaccaactct
     4501 atcagagctt ggttgacggc aatttcgatg atgcagcttg ggcgcagggt cgatgcgacg
     4561 caatcgtccg atccggagcc gggactgtcg ggcgtacaca aatcgcccgc agaagcgcgg
     4621 ccgtctggac cgatggctgt gtagaagtac tcgccgatag tggaaaccga cgccccagca
     4681 ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt cgattccacc gccgccttct
     4741 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     4801 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     4861 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     4921 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
     4981 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     5041 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     5101 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     5161 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     5221 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     5281 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     5341 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     5401 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     5461 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     5521 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     5581 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     5641 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     5701 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     5761 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     5821 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     5881 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     5941 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     6001 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     6061 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     6121 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     6181 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     6241 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     6301 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     6361 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     6421 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     6481 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     6541 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     6601 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     6661 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     6721 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     6781 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     6841 ct
//
