LOCUS       sequence-216265-        6981 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of flag tagged GREB1b.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6981)
  AUTHORS   Haines CN, Braunreiter KM, Mo XM, Burd CJ
  TITLE     GREB1 isoforms regulate proliferation independent of ERalpha 
            co-regulator activities in breast cancer.
  JOURNAL   Endocr Relat Cancer. 2018 Jul;25(7):735-746. doi: 
            10.1530/ERC-17-0496. Epub 2018 Apr 25.
  PUBMED    29695586
REFERENCE   2  (bases 1 to 6981)
  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: 
            "10.1530/ERC-17-0496"; journalName: "Endocr Relat Cancer"; date: 
            "2018-07"; volume: "25"; issue: "7"; pages: "735-746"
COMMENT     Sequence Label: pcDNA-3xflagGREB1b
FEATURES             Location/Qualifiers
     source          1..6981
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      662..671
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             671..736
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(2169..2186)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2175..2399
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2445..2873
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2532..2551)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2742..2763
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2882..2902)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2887..3216
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3067..3202
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3129..3148
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3265..4290
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             3265..4290
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    4420..4541
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4457..4476)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4511..4530
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4590..4606)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4590..4606)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4603..4625)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4614..4630
                     /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(4638..4644,4645..4662,4663..4668))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4683..4704
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4821..4838)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4992..5580)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5072..5091)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5751..6542,6543..6611))
                     /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     6374..6393
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6612..6716)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6791..6810)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 ggccaccatg gactacaaag accatgacgg tgattataaa gatcatgata tcgattacaa
      721 ggatgacgat gacaagctag cgtttaaact taagcttggt accatgggaa attcttacgc
      781 tggacagctg aagacgacac gctttgaaga ggtcttgcac aattccatcg aggcatccct
      841 gcggtccaac aacctggtgc ccaggcccat cttttcccag ctgtacctgg aagctgagca
      901 gcagcttgcc gctctagaag gtggtagccg agtggacaat gaggaagagg aagaagaggg
      961 agaaggaggg ctggaaacaa atggcccccc aaaccctttc cagctgcacc ctctgcctga
     1021 aggatgctgt accacagacg ggttttgcca ggccgggaag gacctgcgcc ttgtctccat
     1081 ttccaacgag cccatggatg tccctgcggg ctttctcctc gtgggggtca agtcccccag
     1141 cctgccggac catctcctgg tgtgcgccgt tgacaagagg ttcttgccag atgacaatgg
     1201 ccacaatgct cttcttggtt tctctgggaa ttgtgttggc tgtggaaaga aaggcttctg
     1261 ttacttcacg gaattctcca atcatataaa tctgaaactg accactcaac ccaagaagca
     1321 gaaacacttg aagtattacc tggtccgtaa tgcacaaggg actctaacca aaggaccttt
     1381 aatctgttgg aaaggctcag agtttagaag ccggcagatc cccgccagta cttgttccag
     1441 ttccctcttc ccagccctgg agagcacggc tgccttcccc agcgagcccg ttcctgggac
     1501 gaaccccagc atcctgatgg gagctcagca ggcaggacca gcttctgatc acccctcact
     1561 aaacgcagca atgggtccgg ctgttttcaa cggcaaagat tccccgaagt gccaacaact
     1621 ggcaaagaat aacctgttgg ccctgccgcg accatcggct ttaggtatct tgtcaaactc
     1681 cgggcccccc aaaaaacgcc acaaagggtg gtctccagaa tctccatcag ctccagatgg
     1741 tggctgcccc caaggtggtg ggaacagagc taagtatgag agcgcaggca tgtcctgcgt
     1801 gccgcaggtt ggcttggtgg gaccagcttc agtcaccttt ccagtggtgg cctctggaga
     1861 accagtgtct gttcctgaca acttgctgaa aatatgcaag gccaagccag tgatatttaa
     1921 aggccatggg aacttccctt acctctgtgg gaacctgaat gacgtcgtgg tcagccccct
     1981 cttgtacacg tgctaccaga attcccagtc tgtctcacgg gcatacgagc agtacggcgc
     2041 ctctgccatc cagcccatct ccgaggagat gcagctcctg cttaccgtct actacctggt
     2101 ccagctggca catcaaatac gaaatccgga cgtataaact agagggcccg tttaaacccg
     2161 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     2221 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     2281 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     2341 caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc
     2401 ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg
     2461 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     2521 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     2581 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     2641 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     2701 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     2761 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     2821 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attaattctg
     2881 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     2941 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     3001 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     3061 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     3121 atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag
     3181 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc
     3241 attttcggat ctgatcagca cgtgatgaaa aagcctgaac tcaccgcgac gtctgtcgag
     3301 aagtttctga tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc ggagggcgaa
     3361 gaatctcgtg ctttcagctt cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc
     3421 tgcgccgatg gtttctacaa agatcgttat gtttatcggc actttgcatc ggccgcgctc
     3481 ccgattccgg aagtgcttga cattggggaa ttcagcgaga gcctgaccta ttgcatctcc
     3541 cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg
     3601 cagccggtcg cggaggccat ggatgcgatc gctgcggccg atcttagcca gacgagcggg
     3661 ttcggcccat tcggaccgca aggaatcggt caatacacta catggcgtga tttcatatgc
     3721 gcgattgctg atccccatgt gtatcactgg caaactgtga tggacgacac cgtcagtgcg
     3781 tccgtcgcgc aggctctcga tgagctgatg ctttgggccg aggactgccc cgaagtccgg
     3841 cacctcgtgc acgcggattt cggctccaac aatgtcctga cggacaatgg ccgcataaca
     3901 gcggtcattg actggagcga ggcgatgttc ggggattccc aatacgaggt cgccaacatc
     3961 ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt cgagcggagg
     4021 catccggagc ttgcaggatc gccgcggctc cgggcgtata tgctccgcat tggtcttgac
     4081 caactctatc agagcttggt tgacggcaat ttcgatgatg cagcttgggc gcagggtcga
     4141 tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc gtacacaaat cgcccgcaga
     4201 agcgcggccg tctggaccga tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc
     4261 cccagcactc gtccgagggc aaaggaatag cacgtgctac gagatttcga ttccaccgcc
     4321 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     4381 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     4441 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     4501 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     4561 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     4621 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     4681 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     4741 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     4801 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     4861 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     4921 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     4981 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     5041 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     5101 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     5161 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     5221 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     5281 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     5341 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     5401 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     5461 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     5521 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     5581 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     5641 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     5701 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     5761 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     5821 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     5881 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     5941 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     6001 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     6061 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     6121 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     6181 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     6241 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     6301 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     6361 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     6421 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     6481 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     6541 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     6601 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     6661 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     6721 tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt
     6781 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
     6841 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
     6901 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
     6961 tacgggccag atatacgcgt t
//
