LOCUS       sequence-440850-        7288 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression vector of flag-tagged GNB1L.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7288)
  AUTHORS   Zhao Y, Tabet D, Rubio Contreras D, Lao L, Kousholt AN, Weile J, 
            Melo H, Hoeg L, Feng S, Cote AG, Lin ZY, Setiaputra D, Jonkers J, 
            Gingras AC, Gomez Herreros F, Roth FP, Durocher D
  TITLE     Genome-scale mapping of DNA damage suppressors through phenotypic 
            CRISPR-Cas9 screens.
  JOURNAL   Mol Cell. 2023 Aug 3;83(15):2792-2809.e9. doi: 
            10.1016/j.molcel.2023.06.025. Epub 2023 Jul 20.
  PUBMED    37478847
REFERENCE   2  (bases 1 to 7288)
  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.2023.06.025"; journalName: "Mol Cell"; date: 
            "2023-08-3- 3"; volume: "83"; issue: "15"; pages: "2792-2809.e9"
COMMENT     Sequence Label: pcDNA5-FRT/T0-EGFP-GNB1L
FEATURES             Location/Qualifiers
     source          1..7288
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      598..1186
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1087..1106
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1340..1357
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1474..1495
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1510..1515,1516..1533,1534..1540)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1548..1564
                     /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     1553..1575
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1572..1588
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1572..1588
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    1637..1758
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1648..1667)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1702..1721
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1888..2910)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     protein_bind    complement(2918..2965)
                     /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)."
     primer_bind     3097..3116
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    3249..3473
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     3462..3479
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(join(5013..5723,5724..5726,5727..5729))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(5013..5723,5724..5726,5727..5729))
                     /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(5055..5076)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     5404..5423
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5663..5684
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5826..5850)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    5860..5878
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5881..5899
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        complement(5901..6104)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(5930..5950)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        6105..6484
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     6656..6675
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        6750..6854
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6855..6923,6924..7288,1..427)
                     /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(7073..7092)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
       61 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
      121 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
      181 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
      241 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
      301 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
      361 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
      421 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
      481 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
      541 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
      601 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
      661 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
      721 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
      781 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
      841 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
      901 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
      961 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     1021 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     1081 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     1141 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     1201 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     1261 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     1321 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg
     1381 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     1441 cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc
     1501 accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata
     1561 acaatttcac acaggaaaca gctatgacca tgattacgcc aagctctagc tagaggtcga
     1621 cggtatacag acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag
     1681 tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata
     1741 agctgcaata aacaagttgg ggtgggcgaa gaactccagc atgagatccc cgcgctggag
     1801 gatcatccag ccggcgtccc ggaaaacgat tccgaagccc aacctttcat agaaggcggc
     1861 ggtggaatcg aaatctcgta gtacgtgcta ttcctttgcc ctcggacgag tgctggggcg
     1921 tcggtttcca ctatcggcga gtacttctac acagccatcg gtccagacgg ccgcgcttct
     1981 gcgggcgatt tgtgtacgcc cgacagtccc ggctccggat cggacgattg cgtcgcatcg
     2041 accctgcgcc caagctgcat catcgaaatt gccgtcaacc aagctctgat agagttggtc
     2101 aagaccaatg cggagcatat acgcccggag ccgcggcgat cctgcaagct ccggatgcct
     2161 ccgctcgaag tagcgcgtct gctgctccat acaagccaac cacggcctcc agaagaagat
     2221 gttggcgacc tcgtattggg aatccccgaa catcgcctcg ctccagtcaa tgaccgctgt
     2281 tatgcggcca ttgtccgtca ggacattgtt ggagccgaaa tccgcgtgca cgaggtgccg
     2341 gacttcgggg cagtcctcgg cccaaagcat cagctcatcg agagcctgcg cgacggacgc
     2401 actgacggtg tcgtccatca cagtttgcca gtgatacaca tggggatcag caatcgcgca
     2461 tatgaaatca cgccatgtag tgtattgacc gattccttgc ggtccgaatg ggccgaaccc
     2521 gctcgtctgg ctaagatcgg ccgcagcgat cgcatccatg gcctccgcga ccggctgcag
     2581 aacagcgggc agttcggttt caggcaggtc ttgcaacgtg acaccctgtg cacggcggga
     2641 gatgcaatag gtcaggctct cgctgaactc cccaatgtca agcacttccg gaatcgggag
     2701 cgcggccgat gcaaagtgcc gataaacata acgatctttg tagaaaccat cggcgcagct
     2761 atttacccgc aggacatatc cacgccctcc tacatcgaag ctgaaagcac gagattcttc
     2821 gccctccgag agctgcatca ggtcggagac gctgtcgaac ttttcgatca gaaacttctc
     2881 gacagacgtc gcggtgagtt caggcttttt ggccaaggaa gttcctatac tttctagaga
     2941 ataggaactt cggaatagga acttctaggt acgtgaacca tcaccctaat caagtttttt
     3001 ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc
     3061 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
     3121 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
     3181 taatgcgccg ctacagggcg cgtggggata ccccctagag ccccagctgg ttctttccgc
     3241 ctcagaagcc atagagccca ccgcatcccc agcatgcctg ctattgtctt cccaatcctc
     3301 ccccttgctg tcctgcccca ccccaccccc cagaatagaa tgacacctac tcagacaatg
     3361 cgatgcaatt tcctcatttt attaggaaag gacagtggga gtggcacctt ccagggtcaa
     3421 ggaaggcacg ggggaggggc aaacaacaga tggctggcaa ctagaaggca cagtcgaggc
     3481 tgatcagcgg gtttaaacgg gccctctaga ctcgagtttt tttttttttt tttttggcaa
     3541 tggaaattta ttataaaata ccctcagctg gcaacacagc aggcccaagc caacgtctcc
     3601 tgcaggcctc ggacggccag ggctctggct ggcccccaga gtcaccgtcc tgtgatgagg
     3661 cactccacaa aaggaaatac caacctcatg aagacagcgg ggactccatg gtccttgggc
     3721 catgacttgc tggtcctcag aggcccttga ggtttcaggc caaggctggg gcctgatgcc
     3781 cacctccctg cccgccctcc tcgtctcccg ggaagggagt gggtgagtca tgcgcgtggg
     3841 tagagtgacc agaggctgat ccgctgatcc ttggagcccg cggccagcaa gccatcggcg
     3901 gtgaaggcca cgcactggac agcggcgctg tggaaggcca gcacggccag tggctgcatc
     3961 gtccgccagt ggaacacgcg gatgcggtgg tcccagcctg cggtggccag gatcttgcga
     4021 tctggccgga tcgtgacctc ggcgatcccg ggattggtga gttcatgagt cccacgcacc
     4081 tgcagggcct gctgccagtc caggctccag acagccagcg ccttccccgc ggagcctgag
     4141 atgcccctgg ccttctggga gtcaaagtca aggtccatga cgggctcctc atggcaggcg
     4201 atgcggctgc acaccttctg ctcagagacg tcccacagga ccaccgatcc atcctcatag
     4261 ccggccagaa ggagtgggcg ggagctgcag tcggcctgcc acagccgcag gcacatgggc
     4321 atgcccagct tggcatctgc cttcggcttc agggcgcaca ctgacgtctt ggagggcatc
     4381 tccagaatct gaacctcgtc gctgcccctc cctggcacgg caagcgtcca gcgtggctgg
     4441 cccccggcca ggatgctgct ccggcagaag cccacactct ccaagcacac ggagtccacg
     4501 acagcgctcc tgccctccgc gaggtcccac aggcacagct tcaggtcccg gccctgactg
     4561 aggagctggc gcccctgggg cagcgtctgc agccaggtca cacactggcc gccgtggcca
     4621 tccagggtgg taaccgctct ccgcgtctgc aggctccaga tgtgtaccag gccactctga
     4681 gaccctgaga agaggagcgg gcgcccctga gcctgggctc cttcgcagaa gtgcagcgca
     4741 tgcaccggtg actgggtgcc tcggaggaca aactgggggt ctggaggtgg cggcgggcag
     4801 ggggccgtca tgctgggcag gatgcagtta cctgaggcta cgcgaatcgc ccgcgaagag
     4861 ggtcctggga tccgtagcca cagaccgtgg gacgggcctc cctctgaggg cgagagtcca
     4921 ggcgccaccc tcttgccgga gtcgggaacc ctcgtgccga attccagcac actggcggcc
     4981 gttactagtg gatccttaat taatggcgcg cccttgtaca gctcgtccat gccgagagtg
     5041 atcccggcgg cggtcacgaa ctccagcagg accatgtgat cgcgcttctc gttggggtct
     5101 ttgctcaggg cggactgggt gctcaggtag tggttgtcgg gcagcagcac ggggccgtcg
     5161 ccgatggggg tgttctgctg gtagtggtcg gcgagctgca cgctgccgtc ctcgatgttg
     5221 tggcggatct tgaagttcac cttgatgccg ttcttctgct tgtcggccat gatatagacg
     5281 ttgtggctgt tgtagttgta ctccagcttg tgccccagga tgttgccgtc ctccttgaag
     5341 tcgatgccct tcagctcgat gcggttcacc agggtgtcgc cctcgaactt cacctcggcg
     5401 cgggtcttgt agttgccgtc gtccttgaag aagatggtgc gctcctggac gtagccttcg
     5461 ggcatggcgg acttgaagaa gtcgtgctgc ttcatgtggt cggggtagcg gctgaagcac
     5521 tgcacgccgt aggtcagggt ggtcacgagg gtgggccagg gcacgggcag cttgccggtg
     5581 gtgcagatga acttcagggt cagcttgccg taggtggcat cgccctcgcc ctcgccggac
     5641 acgctgaact tgtggccgtt tacgtcgccg tccagctcga ccaggatggg caccaccccg
     5701 gtgaacagct cctcgccctt gctcaccatg gtaccaagct taagtttaaa cgctagagtc
     5761 cggaggctgg atcggtcccg gtgtcttcta tggaggtcaa aacagcgtgg atggcgtctc
     5821 caggcgatct gacggttcac taaacgagct cgtcgacgat ctctatcact gatagggaga
     5881 tctctatcac tgatagggag agctctgctt atatagacct cccaccgtac acgcctaccg
     5941 cccatttgcg tcaatggggc ggagttgtta cgacattttg gaaagtcccg ttgattttgg
     6001 tgccaaaaca aactcccatt gacgtcaatg gggtggagac ttggaaatcc ccgtgagtca
     6061 aaccgctatc cacgcccatt gatgtactgc caaaaccgca tcaccatggt aatagcgatg
     6121 actaatacgt agatgtactg ccaagtagga aagtcccata aggtcatgta ctgggcataa
     6181 tgccaggcgg gccatttacc gtcattgacg tcaatagggg gcgtacttgg catatgatac
     6241 acttgatgta ctgccaagtg ggcagtttac cgtaaatact ccacccattg acgtcaatgg
     6301 aaagtcccta ttggcgttac tatgggaaca tacgtcatta ttgacgtcaa tgggcggggg
     6361 tcgttgggcg gtcagccagg cgggccattt accgtaagtt atgtaacgcg gaactccata
     6421 tatgggctat gaactaatga ccccgtaatt gattactatt aataactagt caataatcaa
     6481 tgtcaacgcg tatatctggc ccgtacatcg cgaagcagcg caaaacgcct aaccctaagc
     6541 agattcttca tgcaattgtc ggtcaagcct tgccttgttg tagcttaaat tttgctcgcg
     6601 cactactcag cgacctccaa cacacaagca gggagcagat actggcttaa ctatgcggca
     6661 tcagagcaga ttgtactgag agtgcaccat aggggatcgg gagatctccc gatccgtcga
     6721 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     6781 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     6841 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     6901 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     6961 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     7021 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     7081 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     7141 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     7201 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     7261 ttctgacaac gatcggagga ccgaagga
//
