LOCUS       sequence-420889-        6350 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  LR vector for integration of Chtop-exon into N2a FRT rtTA3 
            expression cells (BioID vector).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6350)
  AUTHORS   Roth JF, Braunschweig U, Wu M, Li JD, Lin ZY, Larsen B, Weatheritt 
            RJ, Gingras AC, Blencowe BJ
  TITLE     Systematic analysis of alternative exon-dependent interactome 
            remodeling reveals multitasking functions of gene regulatory 
            factors.
  JOURNAL   Mol Cell. 2023 Dec 7;83(23):4222-4238.e10. doi: 
            10.1016/j.molcel.2023.10.034.
  PUBMED    38065061
REFERENCE   2  (bases 1 to 6350)
  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: 
            "10.1016/j.molcel.2023.10"; journalName: "Mol Cell"; date: 
            "2023-12-7- 7"; volume: "83"; issue: "23"; pages: "4222-4238.e10"
COMMENT     Sequence Label: 5'-BirA-FLAG Chtop-exon minimal CMV pCDNA5
FEATURES             Location/Qualifiers
     source          1..6350
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    451..572
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(488..507)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     542..561
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(621..637)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(621..637)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(634..656)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    645..661
                     /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(669..675,676..693,694..699))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    714..735
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(852..869)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1023..1611)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1103..1122)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1782..2573,2574..2642))
                     /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     2405..2424
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2643..2747)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2822..2841)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     protein_bind    3040..3058
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3077..3095
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3114..3132
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3151..3169
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     3191..3215
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             join(3348..3698,3699..3701,3702..4310)
                     /codon_start=1
                     /product="promiscuous R118G mutant of the E. coli biotin 
                     protein ligase (Cronan, 2005)"
                     /label=BirA(R118G)
                     /note="also known as BirA* or BioID"
                     /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
                     WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG
                     DACIAEYQQAGRGGRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK
                     LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG
                     WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG
                     DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     CDS             4341..4364
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    4377..4401
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(5014..5038)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(5080..5097)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5086..5310
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5443..5462)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    5594..5641
                     /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             join(5649..6350,1..321)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             join(5649..6350,1..321)
                     /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
                     "
ORIGIN
        1 aggccgtggt tggcttgtat ggagcagcag acgcgctact tcgagcggag gcatccggag
       61 cttgcaggat cgccgcggct ccgggcgtat atgctccgca ttggtcttga ccaactctat
      121 cagagcttgg ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg atgcgacgca
      181 atcgtccgat ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag aagcgcggcc
      241 gtctggaccg atggctgtgt agaagtactc gccgatagtg gaaaccgacg ccccagcact
      301 cgtccgaggg caaaggaata gcacgtacta cgagatttcg attccaccgc cgccttctat
      361 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
      421 gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac
      481 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
      541 tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc
      601 tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
      661 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
      721 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
      781 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
      841 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
      901 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
      961 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     1021 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     1081 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     1141 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     1201 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     1261 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     1321 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     1381 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     1441 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     1501 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     1561 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     1621 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     1681 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     1741 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     1801 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     1861 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     1921 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     1981 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     2041 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     2101 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     2161 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     2221 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     2281 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     2341 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     2401 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     2461 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     2521 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     2581 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     2641 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     2701 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     2761 aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca
     2821 gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg
     2881 aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca
     2941 attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca
     3001 gatatacgcg tctcgaggga tccaagctgg ccgcacgtct ccctatcagt gatagagaag
     3061 tcgacacgtc tcgagctccc tatcagtgat agagaaggta cgtctagaac gtctccctat
     3121 cagtgataga gaagtcgaca cgtctcgagc tccctatcag tgatagagaa ggtaccccct
     3181 atataagcag agctcgttta gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt
     3241 ttgacctcca tagaagacac cgggaccgat ccagcctccg cggaggctgg atcctctagc
     3301 gtttaaactt aagcttggta ccgagctcgg atccagctag cgccaccatg aaggacaaca
     3361 ccgtgcccct gaagctgatc gccctgctgg ccaacggcga gttccacagc ggcgagcagc
     3421 tgggcgagac cctgggcatg agccgggccg ccatcaacaa gcacatccag accctgcggg
     3481 actggggcgt ggacgtgttc accgtgcccg gcaagggcta cagcctgccc gagcccatcc
     3541 agctgctgaa cgccaagcag atcctgggcc agctggacgg cggcagcgtg gccgtgctgc
     3601 ccgtgatcga cagcaccaac cagtacctgc tggaccggat cggcgagctg aagagcggcg
     3661 acgcctgcat cgccgagtac cagcaggccg gccggggcgg ccggggccgg aagtggttca
     3721 gccccttcgg cgccaacctg tacctgagca tgttctggcg gctggagcag ggccccgccg
     3781 ccgccatcgg cctgagcctg gtgatcggca tcgtgatggc cgaggtgctg cggaagctgg
     3841 gcgccgacaa ggtgcgggtg aagtggccca acgacctgta cctgcaggac cggaagctgg
     3901 ccggcatcct ggtggagctg accggcaaga ccggcgacgc cgcccagatc gtgatcggcg
     3961 ccggcatcaa catggccatg cggcgggtgg aggagagcgt ggtgaaccag ggctggatca
     4021 ccctgcagga ggccggcatc aacctggacc ggaacaccct ggccgccatg ctgatccggg
     4081 agctgcgggc cgccctggag ctgttcgagc aggagggcct ggccccctac ctgagccggt
     4141 gggagaagct ggacaacttc atcaaccggc ccgtgaagct gatcatcggc gacaaggaga
     4201 tcttcggcat cagccggggc atcgacaagc agggcgccct gctgctggag caggacggca
     4261 tcatcaagcc ctggatgggc ggcgagatca gcctgcggag cgccgagaag gggggatctg
     4321 gccccggcgg cggcgcgcca gactacaaag acgatgacga taaactaggc gcgcccacaa
     4381 gtttgtacaa aaaagttggc accgctgcac agtcagcgcc gaaagttgtg ctaaaaagca
     4441 ccaccaagat gtctctaaat gagcgcttta ctaatatgct gaagaacaaa cagccgatgc
     4501 cagtgaatat tcgggcttcg atgcagcagc agcagcagct agccagtgcc agaaacagaa
     4561 gactggccca gcagatggag aatagaccct ctgtccaggc agcattaaaa cttaagcaga
     4621 agagcttaaa gcagcgcctg ggtaagagta atatccaggc acggttaggc cgacccatag
     4681 gtgccctggc caggggagca attggaggaa gaggcctacc cataatccag agaggcttgc
     4741 cccgaggagg actacgtggg ggacgtgcta ccagaaccct gcttaggggt gggatgtcgc
     4801 tccgaggtcg gggtatgata ggtcggggaa gagggggctt tggaggcaga ggccgaggtc
     4861 gtggccgagg gagaggtgcc ctcactcgcc ctgtattgac caaggagcag ctggacaacc
     4921 aattggatgc atacatgtcg aaaactaaag gacacctgga tgctgaattg gatgcctaca
     4981 tggcacagac agatcctgaa accaatgatt gatacccaac tttcttgtac aaagtggtgt
     5041 gactcgagtc tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt
     5101 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     5161 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     5221 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     5281 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
     5341 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     5401 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     5461 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     5521 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     5581 gttcacgtac ctagaagttc ctattccgaa gttcctattc tctagaaagt ataggaactt
     5641 ccttggccaa aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg atcgaaaagt
     5701 tcgacagcgt ctccgacctg atgcagctct cggagggcga agaatctcgt gctttcagct
     5761 tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat ggtttctaca
     5821 aagatcgtta tgtttatcgg cactttgcat cggccgcgct cccgattccg gaagtgcttg
     5881 acattgggga attcagcgag agcctgacct attgcatctc ccgccgtgca cagggtgtca
     5941 cgttgcaaga cctgcctgaa accgaactgc ccgctgttct gcagccggtc gcggaggcca
     6001 tggatgcgat cgctgcggcc gatcttagcc agacgagcgg gttcggccca ttcggaccgc
     6061 aaggaatcgg tcaatacact acatggcgtg atttcatatg cgcgattgct gatccccatg
     6121 tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg caggctctcg
     6181 atgagctgat gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg cacgcggatt
     6241 tcggctccaa caatgtcctg acggacaatg gccgcataac agcggtcatt gactggagcg
     6301 aggcgatgtt cggggattcc caatacgagg tcgccaacat cttcttctgg
//
