LOCUS       sequence-454030-        5140 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes constitutive expression of mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5140)
  AUTHORS   McNamara HM, Solley SC, Adamson B, Chan MM, Toettcher JE
  TITLE     Recording morphogen signals reveals mechanisms underlying gastruloid
            symmetry breaking.
  JOURNAL   Nat Cell Biol. 2024 Oct 2. doi: 10.1038/s41556-024-01521-9.
  PUBMED    39358450
REFERENCE   2  (bases 1 to 5140)
  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: "Nat Cell 
            Biol. 2024 Oct 2. doi: 10.1038/s41556-024-01521-9."
COMMENT     Sequence Label: pHM028
FEATURES             Location/Qualifiers
     source          1..5140
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   23..57
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        432..931
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(467..490)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     864..883
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             952..1662
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1097..1115)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1365..1384
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    1763..2351
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1816..1836)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2234..2245)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     repeat_region   2563..2625
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(2737..2756)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2856..2878
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2916..2934)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3002..3106
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3107..3175,3176..3967)
                     /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(3325..3344)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4138..4726
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4627..4646
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4880..4897
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5014..5035
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5050..5055,5056..5073,5074..5080)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5088..5104
                     /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     5093..5115
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5112..5128
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5112..5128
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ggggcgcgcc attctagatt aaccctagaa agatagtctg cgtaaaattg acgcatgcat
       61 tcttgaaata ttgctctctc tttctaaata gcgcgaatcc gtcgctgtgc atttaggaca
      121 tctcagtcgc cgcttggagc tcccgtgagg cgtgcttgtc aatgcggtaa gtgtcactga
      181 ttttgaacta taacgaccgc gtgagtcaaa atgacgcatg attatctttt acgtgacttt
      241 taagatttaa ctcatacgat aattatattg ttatttcatg ttctacttac gtgataactt
      301 attatatata tattttcttg ttatagatat caactagaat gctagcatgg gcccagatct
      361 ccgaggggac ccgacaggcc cgcagggaca gcagagatcc agtttggatc gataagcttg
      421 atatcgaatt ggggtagggg aggcgctttt cccaaggcag tctggagcat gcgctttagc
      481 agccccgctg ggcacttggc gctacacaag tggcctctgg cctcgcacac attccacatc
      541 caccggtagg cgccaaccgg ctccgttctt tggtggcccc ttcgcgccac cttctactcc
      601 tcccctagtc aggaagttcc cccccgcccc gcagctcgcg tcgtgcagga cgtgacaaat
      661 ggaagtagca cgtctcacta gtctcgtgca gatggacagc accgctgagc aatggaagcg
      721 ggtaggcctt tggggcagcg gccaatagca gctttgctcc ttcgctttct gggctcagag
      781 gctgggaagg ggtgggtccg ggggcgggct caggggcggg ctcaggggcg gggcgggcgc
      841 ccgaaggtcc tccggaggcc cggcattctg cacgcttcaa aagcgcacgt ctgccgcgct
      901 gttctcctct tcctcatctc cgggcctttc gacctgcagc ccaagcttac catggtgagc
      961 aagggcgagg aggataacat ggccatcatc aaggagttca tgcgcttcaa ggtgcacatg
     1021 gagggctccg tgaacggcca cgagttcgag atcgagggcg agggcgaggg ccgcccctac
     1081 gagggcaccc agaccgccaa gctgaaggtg accaagggtg gccccctgcc cttcgcctgg
     1141 gacatcctgt cccctcagtt catgtacggc tccaaggcct acgtgaagca ccccgccgac
     1201 atccccgact acttgaagct gtccttcccc gagggcttca agtgggagcg cgtgatgaac
     1261 ttcgaggacg gcggcgtggt gaccgtgacc caggactcct ccctgcagga cggcgagttc
     1321 atctacaagg tgaagctgcg cggcaccaac ttcccctccg acggccccgt aatgcagaag
     1381 aagaccatgg gctgggaggc ctcctccgag cggatgtacc ccgaggacgg cgccctgaag
     1441 ggcgagatca agcagaggct gaagctgaag gacggcggcc actacgacgc tgaggtcaag
     1501 accacctaca aggccaagaa gcccgtgcag ctgcccggcg cctacaacgt caacatcaag
     1561 ttggacatca cctcccacaa cgaggactac accatcgtgg aacagtacga acgcgccgag
     1621 ggccgccact ccaccggcgg catggacgag ctgtacaagt aacagatcgg cgtgatggaa
     1681 gagtaaagcg gccgcgactc tagagtcgac ctgcaggcat gcaagcttgc aggcatgcaa
     1741 gcttgatatc aagcttatcg ataatcaacc tctggattac aaaatttgtg aaagattgac
     1801 tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt
     1861 gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt
     1921 gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt
     1981 gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg
     2041 gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg
     2101 ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaaatc
     2161 atcgtccttt ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt
     2221 ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc
     2281 tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc
     2341 cgcctccccg ccgaaaggag cgcacgaccc catgcatcga tactagttaa aagttttgtt
     2401 actttataga agaaattttg agtttttgtt tttttttaat aaataaataa acataaataa
     2461 attgtttgtt gaatttatta ttagtatgta agtgtaaata taataaaact taatatctat
     2521 tcaaattaat aaataaacct cgatatacag accgataaaa cacatgcgtc aattttacgc
     2581 atgattatct ttaacgtacg tcacaatatg attatctttc tagggttaat ctagtatacg
     2641 cgtatgcggc cgcttaatta atccatggcg cctgatgcgg tattttctcc ttacgcatct
     2701 gtgcggtatt tcacaccgca tatggtgcac tctcagtaca atctgctctg atgccgcata
     2761 gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct
     2821 cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt
     2881 ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc tatttttata
     2941 ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc ggggaaatgt
     3001 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     3061 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
     3121 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     3181 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
     3241 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
     3301 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
     3361 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
     3421 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     3481 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     3541 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     3601 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     3661 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     3721 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     3781 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     3841 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     3901 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     3961 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     4021 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     4081 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     4141 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     4201 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     4261 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     4321 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     4381 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     4441 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     4501 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     4561 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     4621 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     4681 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     4741 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     4801 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     4861 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg
     4921 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     4981 cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc
     5041 accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata
     5101 acaatttcac acaggaaaca gctatgacca tgattacgcc
//
