LOCUS       sequence-349768-        6481 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression vector encoding CYYR1 and EmGFP-Bsd.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6481)
  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 6481)
  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: CYYR1_pcDNA6.2/EmGFP-Bsd
FEATURES             Location/Qualifiers
     source          1..6481
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        62..109
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             join(128..130,131..133,134..844)
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(173..194)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(434..453)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     781..802
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             845..1240
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="AKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTGV
                     NVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGIK
                     AIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    1398..1519
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1435..1454)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1489..1508
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1568..1584)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1568..1584)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1581..1603)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1592..1608
                     /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(1616..1622,1623..1640,1641..1646))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1661..1682
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1799..1816)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1970..2558)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2050..2069)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2729..3520,3521..3589))
                     /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     3352..3371
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3590..3694)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3769..3788)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3960..4339
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4340..4543
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4494..4514
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4588..4607
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4588..4606
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4636..4660
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(5162..5186)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             5212..5253
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(5293..5312)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5337..5385
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5587..6015
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5674..5693)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5884..5905
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        join(6030..6481,1..48)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(6065..6088)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     6462..6481
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
ORIGIN
        1 cgcacgtctg ccgcgctgtt ctcctcttcc tcatctccgg gcctttcgac tctagacacg
       61 tgttgacaat taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa
      121 ctaaaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
      181 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
      241 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
      301 gcccaccctc gtgaccacct tcacctacgg cgtgcagtgc ttcgcccgct accccgacca
      361 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
      421 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
      481 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
      541 ggggcacaag ctggagtaca actacaacag ccacaaggtc tatatcaccg ccgacaagca
      601 gaagaacggc atcaaggtga acttcaagac ccgccacaac atcgaggacg gcagcgtgca
      661 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
      721 caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca
      781 catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta
      841 caaggccaag cctttgtctc aagaagaatc caccctcatt gaaagagcaa cggctacaat
      901 caacagcatc cccatctctg aagactacag cgtcgccagc gcagctctct ctagcgacgg
      961 ccgcatcttc actggtgtca atgtatatca ttttactggg ggaccttgtg cagaactcgt
     1021 ggtgctgggc actgctgctg ctgcggcagc tggcaacctg acttgtatcg tcgcgatcgg
     1081 aaatgagaac aggggcatct tgagcccctg cggacggtgc cgacaggtgc ttctcgatct
     1141 gcatcctggg atcaaagcca tagtgaagga cagtgatgga cagccgacgg cagttgggat
     1201 tcgtgaattg ctgccctctg gttatgtgtg ggagggctaa gcacttcgtg gccgaggagc
     1261 aggactgaca cgtgctacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     1321 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     1381 agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     1441 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     1501 aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt
     1561 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     1621 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     1681 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     1741 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     1801 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     1861 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     1921 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     1981 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     2041 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     2101 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     2161 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     2221 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     2281 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     2341 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     2401 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     2461 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     2521 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     2581 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     2641 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     2701 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     2761 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     2821 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     2881 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     2941 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     3001 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     3061 cacgctcgtc gtttggtatg gcttcattca gcttcggttc ccaacgatca aggcgagtta
     3121 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     3181 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     3241 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     3301 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     3361 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     3421 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     3481 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     3541 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     3601 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     3661 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     3721 acgtcgacgg atcgggagat ctcccgatcc cctatggtgc actctcagta caatctgctc
     3781 tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta
     3841 gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa
     3901 tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg
     3961 acattgatta ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc
     4021 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
     4081 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
     4141 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
     4201 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
     4261 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
     4321 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
     4381 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
     4441 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
     4501 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctctggc taactagaga
     4561 acccactgct tactggctta tcgaaattaa tacgactcac tatagggaga cccaagctgg
     4621 ctagttaagc tatcaacaag tttgtacaaa aaagcaggct ccgcggccgc ccccttcacc
     4681 atggacgctc cgaggctacc cgtgcgtcca ggggtcttgc ttccgaagtt ggtcctgctc
     4741 tttgtctacg cagatgattg ccttgctcag tgtggcaaag attgcaaatc ttactgctgt
     4801 gatggaacca cgccctactg ttgctcctac tacgcttata ttgggaatat cctctcgggc
     4861 actgcaattg cgggcattgt ttttggaata gtatttatca tgggggtcat tgctgggatt
     4921 gccatatgca tctgcatgtg catgaagaac cacagggcga cccgcgtggg catcctcagg
     4981 acgactcaca tcaacaccgt ctcctcctat cctggaccac caccctacgg tcacgaccac
     5041 gagatggaat actgtgcaga cttgcctcct ccatactccc ccaccccaca gggtccagca
     5101 cagcgttctc caccccctcc ttatcctgga aacgcaagga aataaaaggg tgggcgcgcc
     5161 gacccagctt tcttgtacaa agtggttgat ctagagggcc cgcggttcga aggtaagcct
     5221 atccctaacc ctctcctcgg tctcgattct acgcgtaccg gttagtaatg agtttaaacg
     5281 ggggaggcta actgaaacac ggaaggagac aataccggaa ggaacccgcg ctatgacggc
     5341 aataaaaaga cagaataaaa cgcacgggtg ttgggtcgtt tgttcataaa cgcggggttc
     5401 ggtcccaggg ctggcactct gtcgataccc caccgagacc ccattggggc caatacgccc
     5461 gcgtttcttc cttttcccca ccccaccccc caagttcggg tgaaggccca gggctcgcag
     5521 ccaacgtcgg ggcggcaggc cctgccatag cagatctgcg cagctggggc tctagggggt
     5581 atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     5641 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     5701 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     5761 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     5821 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     5881 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     5941 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     6001 aatttaacgc gaattatcct cgagctaccg ggtaggggag gcgcttttcc caaggcagtc
     6061 tggagcatgc gctttagcag ccccgctggg cacttggcgc tacacaagtg gcctctggcc
     6121 tcgcacacat tccacatcca ccggtaggcg ccaaccggct ccgttctttg gtggcccctt
     6181 cgcgccacct tctactcctc ccctagtcag gaagttcccc cccgccccgc agctcgcgtc
     6241 gtgcaggacg tgacaaatgg aagtagcacg tctcactagt ctcgtgcaga tggacagcac
     6301 cgctgagcaa tggaagcggg taggcctttg gggcagcggc caatagcagc tttgctcctt
     6361 cgctttctgg gctcagaggc tgggaagggg tgggtccggg ggcgggctca ggggcgggct
     6421 caggggcggg gcgggcgccc gaaggtcctc cggaggcccg gcattctgca cgcttcaaaa
     6481 g
//
