LOCUS       sequence-349734-        7426 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression vector encoding RUNX1 and EmGFP-Bsd.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7426)
  AUTHORS   Moyer AJ, Fernandez FX, Li Y, Klinedinst DK, Florea LD, Kazuki Y, 
            Oshimura M, Reeves RH
  TITLE     Overexpression screen of chromosome 21 genes reveals modulators of 
            Sonic hedgehog signaling relevant to Down syndrome.
  JOURNAL   Dis Model Mech. 2023 Apr 1;16(4):dmm049712. doi: 10.1242/dmm.049712.
            Epub 2023 Apr 13.
  PUBMED    36995257
REFERENCE   2  (bases 1 to 7426)
  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: "Dis Model 
            Mech."; date: "2023-04-1"; volume: "16(4):dmm049712. doi"; pages: " 
            10.1242/dmm.049712. Epub 2023 Apr 13"
COMMENT     Sequence Label: RUNX1_pcDNA6.2/EmGFP-Bsd
FEATURES             Location/Qualifiers
     source          1..7426
                     /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"
     protein_bind    677..701
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2148..2172)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2198..2239
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(2279..2298)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2323..2371
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2573..3001
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2660..2679)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2870..2891
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3016..3515
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3051..3074)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3448..3467
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     promoter        3529..3576
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             join(3595..3597,3598..3600,3601..4311)
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3640..3661)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3901..3920)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4248..4269
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             4312..4707
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="AKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTGV
                     NVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGIK
                     AIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    4865..4986
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4902..4921)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4956..4975
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5035..5051)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5035..5051)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5048..5070)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5059..5075
                     /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(5083..5089,5090..5107,5108..5113))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5128..5149
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5266..5283)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5437..6025)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5517..5536)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6196..6987,6988..7056))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPKLNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6819..6838
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7057..7161)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7236..7255)
                     /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 gctagttaag ctatcaacaa gtttgtacaa aaaagcaggc accatggctt cagacagcat
      721 atttgagtca tttccttcgt acccacagtg cttcatgaga gaatgcatac ttggaatgaa
      781 tccttctaga gacgtccacg atgccagcac gagccgccgc ttcacgccgc cttccaccgc
      841 gctgagccca ggcaagatga gcgaggcgtt gccgctgggc gccccggacg ccggcgctgc
      901 cctggccggc aagctgagga gcggcgaccg cagcatggtg gaggtgctgg ccgaccaccc
      961 gggcgagctg gtgcgcaccg acagccccaa cttcctctgc tccgtgctgc ctacgcactg
     1021 gcgctgcaac aagaccctgc ccatcgcttt caaggtggtg gccctagggg atgttccaga
     1081 tggcactctg gtcactgtga tggctggcaa tgatgaaaac tactcggctg agctgagaaa
     1141 tgctaccgca gccatgaaga accaggttgc aagatttaat gacctcaggt ttgtcggtcg
     1201 aagtggaaga gggaaaagct tcactctgac catcactgtc ttcacaaacc caccgcaagt
     1261 cgccacctac cacagagcca tcaaaatcac agtggatggg ccccgagaac ctcgaagaca
     1321 tcggcagaaa ctagatgatc agaccaagcc cgggagcttg tccttttccg agcggctcag
     1381 tgaactggag cagctgcggc gcacagccat gagggtcagc ccacaccacc cagcccccac
     1441 gcccaaccct cgtgcctccc tgaaccactc cactgccttt aaccctcagc ctcagagtca
     1501 gatgcaggat acaaggcaga tccaaccatc cccaccgtgg tcctacgatc agtcctacca
     1561 atacctggga tccattgcct ctccttctgt gcacccagca acgcccattt cacctggacg
     1621 tgccagcggc atgacaaccc tctctgcaga actttccagt cgactctcaa cggcacccga
     1681 cctgacagcg ttcagcgacc cgcgccagtt ccccgcgctg ccctccatct ccgacccccg
     1741 catgcactat ccaggcgcct tcacctactc cccgacgccg gtcacctcgg gcatcggcat
     1801 cggcatgtcg gccatgggct cggccacgcg ctaccacacc tacctgccgc cgccctaccc
     1861 cggctcgtcg caagcgcagg gaggcccgtt ccaagccagc tcgccctcct accacctgta
     1921 ctacggcgcc tcggccggct cctaccagtt ctccatggtg ggcggcgagc gctcgccgcc
     1981 gcgcatcctg ccgccctgca ccaacgcctc caccggctcc gcgctgctca accccagcct
     2041 cccgaaccag agcgacgtgg tggaggccga gggcagccac agcaactccc ccaccaacat
     2101 ggcgccctcc gcgcgcctgg aggaggccgt gtggaggccc tactgaaacc cagctttctt
     2161 gtacaaagtg gttgatctag agggcccgcg gttcgaaggt aagcctatcc ctaaccctct
     2221 cctcggtctc gattctacgc gtaccggtta gtaatgagtt taaacggggg aggctaactg
     2281 aaacacggaa ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga
     2341 ataaaacgca cgggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg
     2401 cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt
     2461 tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg
     2521 gcaggccctg ccatagcaga tctgcgcagc tggggctcta gggggtatcc ccacgcgccc
     2581 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     2641 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     2701 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     2761 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     2821 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     2881 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     2941 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     3001 tatcctcgag ctaccgggta ggggaggcgc ttttcccaag gcagtctgga gcatgcgctt
     3061 tagcagcccc gctgggcact tggcgctaca caagtggcct ctggcctcgc acacattcca
     3121 catccaccgg taggcgccaa ccggctccgt tctttggtgg ccccttcgcg ccaccttcta
     3181 ctcctcccct agtcaggaag ttcccccccg ccccgcagct cgcgtcgtgc aggacgtgac
     3241 aaatggaagt agcacgtctc actagtctcg tgcagatgga cagcaccgct gagcaatgga
     3301 agcgggtagg cctttggggc agcggccaat agcagctttg ctccttcgct ttctgggctc
     3361 agaggctggg aaggggtggg tccgggggcg ggctcagggg cgggctcagg ggcggggcgg
     3421 gcgcccgaag gtcctccgga ggcccggcat tctgcacgct tcaaaagcgc acgtctgccg
     3481 cgctgttctc ctcttcctca tctccgggcc tttcgactct agacacgtgt tgacaattaa
     3541 tcatcggcat agtatatcgg catagtataa tacgacaagg tgaggaacta aaccatggtg
     3601 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     3661 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     3721 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     3781 accaccttca cctacggcgt gcagtgcttc gcccgctacc ccgaccacat gaagcagcac
     3841 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     3901 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     3961 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     4021 gagtacaact acaacagcca caaggtctat atcaccgccg acaagcagaa gaacggcatc
     4081 aaggtgaact tcaagacccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
     4141 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     4201 agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     4261 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa ggccaagcct
     4321 ttgtctcaag aagaatccac cctcattgaa agagcaacgg ctacaatcaa cagcatcccc
     4381 atctctgaag actacagcgt cgccagcgca gctctctcta gcgacggccg catcttcact
     4441 ggtgtcaatg tatatcattt tactggggga ccttgtgcag aactcgtggt gctgggcact
     4501 gctgctgctg cggcagctgg caacctgact tgtatcgtcg cgatcggaaa tgagaacagg
     4561 ggcatcttga gcccctgcgg acggtgccga caggtgcttc tcgatctgca tcctgggatc
     4621 aaagccatag tgaaggacag tgatggacag ccgacggcag ttgggattcg tgaattgctg
     4681 ccctctggtt atgtgtggga gggctaagca cttcgtggcc gaggagcagg actgacacgt
     4741 gctacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     4801 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     4861 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     4921 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     4981 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     5041 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     5101 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     5161 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     5221 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     5281 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     5341 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     5401 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     5461 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     5521 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     5581 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     5641 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     5701 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     5761 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     5821 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     5881 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     5941 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     6001 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     6061 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     6121 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     6181 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     6241 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     6301 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     6361 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     6421 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     6481 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     6541 tggtatggct tcattcagct tcggttccca acgatcaagg cgagttacat gatcccccat
     6601 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     6661 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     6721 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     6781 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     6841 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     6901 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     6961 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     7021 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     7081 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     7141 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc
     7201 gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag
     7261 ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa
     7321 atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt
     7381 aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgtt
//
