LOCUS       sequence-320733-        5446 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive expression of DsRed-Express2-ZF21 under the CMV 
            promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5446)
  TITLE     Leonard Lab COMET plasmids
REFERENCE   2  (bases 1 to 5446)
  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"
COMMENT     Sequence Label: pPD708 DsRed-Express2-ZF21
FEATURES             Location/Qualifiers
     source          1..5446
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      901..910
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             910..975
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             982..1002
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1045..1719
                     /codon_start=1
                     /product="noncytotoxic tetrameric variant of DsRed 
                     fluorescent protein (Strack et al., 2008)"
                     /label=DsRed-Express2
                     /note="mammalian codon-optimized"
                     /translation="MDSTENVIKPFMRFKVHMEGSVNGHEFEIEGEGEGKPYEGTQTAK
                     LQVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGTFIYHVKFIGVNFPSDGPVMQKKTLGWEPSTERLYPRDGVLKGEIHK
                     ALKLKGGGHYLVEFKSIYMAKKPVKLPGYYYVDSKLDITSHNEDYTVVEQYERAEARHH
                     LFQ"
     primer_bind     complement(1225..1245)
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
     primer_bind     1637..1660
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(2076..2093)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2082..2306
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2352..2780
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2439..2458)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2649..2670
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2803..2999
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2850..2985
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2912..2931
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    3119..3240
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3156..3175)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3210..3229
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3289..3305)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3289..3305)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3302..3324)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3313..3329
                     /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(3337..3343,3344..3361,3362..3367))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3382..3403
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3520..3537)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3691..4279)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3771..3790)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4450..5241,5242..5310))
                     /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     5073..5092
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5311..5415)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagtcccaa gctggctagc
      901 gccaccatgg attataagga tcacgatggc gactacaagg accacgatat tgactacaaa
      961 gacgatgacg ataaaatggc tcctaagaaa aagcgcaaag tcggtatcca tggcgttccc
     1021 ggatcaggct ccggaagtgg ctccatggat agcactgaga acgtcatcaa gcccttcatg
     1081 cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag
     1141 ggcgagggca agccctacga gggcacccag accgccaagc tccaggtgac caagggcggc
     1201 cccctgccct tcgcctggga catcctgtcc ccccagttcc agtacggctc caaggtgtac
     1261 gtgaagcacc ccgccgacat ccccgactac aagaagctgt ccttccccga gggcttcaag
     1321 tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc
     1381 ctccaggacg gcaccttcat ctaccacgtg aagttcatcg gcgtgaactt cccctccgac
     1441 ggccccgtaa tgcagaagaa aactctgggc tgggagccct ccaccgagcg cctgtacccc
     1501 cgcgacggcg tgctgaaggg cgagatccac aaggcgctga agctgaaggg cggcggccac
     1561 tacctggtgg agttcaagtc aatctacatg gccaagaagc ccgtgaagct gcccggctac
     1621 tactacgtgg actccaagct ggacatcacc tcccacaacg aggactacac cgtggtggag
     1681 cagtacgagc gcgccgaggc ccgccaccac ctgttccagt ctggttctgg ttctggttca
     1741 ggatccggta ccggtggagc tagacccggg gagcgcccct tccagtgtcg catttgcatg
     1801 cggaactttt cgagaaattt cattttgcaa agacataccc gtactcatac cggtgaaaaa
     1861 ccgtttcagt gtcggatctg tatgcgaaat ttctcccaag gtggtaattt ggttagacat
     1921 ctacgtacgc acaccggcga gaagccattc caatgccgaa tatgcatgcg caacttcagt
     1981 caacaaactg gtttgaatgt tcacctaaaa acccacctga gaggatctta agcggccgct
     2041 cgagtctaga gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca
     2101 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     2161 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
     2221 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagtca atagcaggca
     2281 tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag
     2341 ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     2401 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     2461 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     2521 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     2581 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     2641 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     2701 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     2761 acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttaggctc aattagtcag
     2821 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     2881 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     2941 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     3001 agctcccggg agcttgtata tccattttcg gaggttgggc ttcggaatcg ttttccggga
     3061 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     3121 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     3181 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     3241 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt
     3301 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     3361 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     3421 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     3481 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     3541 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     3601 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     3661 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     3721 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     3781 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     3841 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     3901 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     3961 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     4021 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     4081 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     4141 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     4201 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     4261 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     4321 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     4381 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     4441 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     4501 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     4561 atctggcccc agtgctgcaa tgataccgcg agatccacgc tcaccggctc cagatttatc
     4621 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     4681 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     4741 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     4801 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     4861 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     4921 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     4981 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     5041 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     5101 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     5161 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     5221 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     5281 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     5341 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     5401 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
