LOCUS       sequence-214111-        6592 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses both zinc alpha-2 glycoprotein (ZAG) and dTomato (dT) 
            reporter, where the fluorescent dT reporter is fused with the ZAG 
            protein..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6592)
  AUTHORS   Elattar S, Dimri M, Satyanarayana A
  TITLE     The tumor secretory factor ZAG promotes white adipose tissue 
            browning and energy wasting.
  JOURNAL   FASEB J. 2018 Mar 23:fj201701465RR. doi: 10.1096/fj.201701465RR.
  PUBMED    29570397
REFERENCE   2  (bases 1 to 6592)
  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: "FASEB J. 
            2018 Mar 23:fj201701465RR. doi: 10.1096/fj.201701465RR."
COMMENT     Sequence Label: pRP(Exp)-CMV> ORF_924bp (Azgp1):dTomato:IRES:Puro
FEATURES             Location/Qualifiers
     source          1..6592
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    1..49
                     /note="synthetic polyadenylation signal"
     misc_feature    63..154
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     103..122
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     protein_bind    190..210
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     enhancer        271..574
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        575..778
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     729..749
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    799..823
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      824..833
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1751..2452
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLYGMDELYK"
     primer_bind     complement(1884..1902)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2152..2171
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2346..2369
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     protein_bind    complement(2456..2480)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    join(2481..3055,3056..3058,3059..3067)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2648..2665)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2875..2894
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3068..3667
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3068..3087)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3564..3584
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     protein_bind    complement(3668..3688)
                     /gene="mutant version of attB"
                     /label=attB3
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     polyA_signal    3716..3837
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3727..3746)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3781..3800
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4085..4102)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4256..4844)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4336..4355)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5015..5806,5807..5875))
                     /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     5638..5657
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5876..5980)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6007..6462
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6094..6113)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6304..6325
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtga atcgatagta
       61 ctaacatacg ctctccatca aaacaaaacg aaacaaaaca aactagcaaa ataggctgtc
      121 cccagtgcaa gtgcaggtgc cagaacattt ctctatcgat aggtaccgag ctcttacgcg
      181 tgctagcatc aactttgtat agaaaagttg tagttattaa tagtaatcaa ttacggggtc
      241 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
      301 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
      361 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
      421 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg
      481 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca
      541 gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa
      601 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa
      661 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc
      721 cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctgg
      781 tttagtgaac cgtcagatcc aagtttgtac aaaaaagcag gctgccacca tggtgcctgt
      841 cctgctgtcc ctgcctctcc ttctgggtcc tgcagtcttt caggagactg ggtcttatta
      901 tctgaccttt ctctacaccg ggttgtccag acccagcaaa ggttttccga ggtttcaagc
      961 cactgcattt ctcaatgacc aggccttctt ccactacaac agcaacagcg ggaaggcaga
     1021 gcctgtggga ccttggagcc aggtggaagg aatggaggac tgggagaagg aaagccagct
     1081 tcagagggcc agggaggaga tcttccttgt gaccctgaaa gacatcatgg actattacaa
     1141 ggacactaca gggtctcaca cctttcaggg aatgtttggt tgcgagatca caaataacag
     1201 aagtagtgga gctgtctgga ggtatgccta cgacggagag gatttcatcg aattcaacaa
     1261 agaaatccca gcttggatcc ccttagaccc agcagctgca aacaccaagc taaagtggga
     1321 agcagaaaag gtctacgtgc agcgagccaa ggcataccta gaggaggagt gtcctgaaat
     1381 gctgaagagg tacctgaact acagtcgatc tcacctggac cgaatagatc ctcccactgt
     1441 gacaatcacc agccgtgtga tcccaggagg aaacagaata ttcaaatgcc tggcctatgg
     1501 cttctaccca caaagaatta gtctgcactg gaacaaggcc aacaagaagc tagcatttga
     1561 accagaaaga ggtgtttttc ccaatggaaa tggcacttac ctctcctggg cagaggtgga
     1621 agtctcccca caggacatag accccttctt ctgcctcata gatcacaggg ggttttccca
     1681 atctctctcg gtgcagtggg ataggacaag aaaagtaaag gatgaaaaca atgttgtagc
     1741 tcagcctcag atggtgagca agggcgagga ggtcatcaaa gagttcatgc gcttcaaggt
     1801 gcgcatggag ggctccatga acggccacga gttcgagatc gagggcgagg gcgagggccg
     1861 cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggcggcc ccctgccctt
     1921 cgcctgggac atcctgtccc cccagttcat gtacggctcc aaggcgtacg tgaagcaccc
     1981 cgccgacatc cccgattaca agaagctgtc cttccccgag ggcttcaagt gggagcgcgt
     2041 gatgaacttc gaggacggcg gtctggtgac cgtgacccag gactcctccc tgcaggacgg
     2101 cacgctgatc tacaaggtga agatgcgcgg caccaacttc ccccccgacg gccccgtaat
     2161 gcagaagaag accatgggct gggaggcctc caccgagcgc ctgtaccccc gcgacggcgt
     2221 gctgaagggc gagatccacc aggccctgaa gctgaaggac ggcggccact acctggtgga
     2281 gttcaagacc atctacatgg ccaagaagcc cgtgcaactg cccggctact actacgtgga
     2341 caccaagctg gacatcacct cccacaacga ggactacacc atcgtggaac agtacgagcg
     2401 ctccgagggc cgccaccacc tgttcctgta cggcatggac gagctgtaca agtaaaccca
     2461 gctttcttgt acaaagtggg cccctctccc tccccccccc ctaacgttac tggccgaagc
     2521 cgcttggaat aaggccggtg tgcgtttgtc tatatgttat tttccaccat attgccgtct
     2581 tttggcaatg tgagggcccg gaaacctggc cctgtcttct tgacgagcat tcctaggggt
     2641 ctttcccctc tcgccaaagg aatgcaaggt ctgttgaatg tcgtgaagga agcagttcct
     2701 ctggaagctt cttgaagaca aacaacgtct gtagcgaccc tttgcaggca gcggaacccc
     2761 ccacctggcg acaggtgcct ctgcggccaa aagccacgtg tataagatac acctgcaaag
     2821 gcggcacaac cccagtgcca cgttgtgagt tggatagttg tggaaagagt caaatggctc
     2881 tcctcaagcg tattcaacaa ggggctgaag gatgcccaga aggtacccca ttgtatggga
     2941 tctgatctgg ggcctcggtg cacatgcttt acatgtgttt agtcgaggtt aaaaaaacgt
     3001 ctaggccccc cgaaccacgg ggacgtggtt ttcctttgaa aaacacgatg ataatatggc
     3061 cacaaccatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag
     3121 ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga
     3181 tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg
     3241 gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac
     3301 gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt
     3361 gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc
     3421 caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg
     3481 tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc
     3541 cttcctggag acctccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt
     3601 caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg
     3661 tgcctgacaa ctttattata catagttgat ggccggccgc ttcgagcaga catgataaga
     3721 tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt
     3781 gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa acaagttaac
     3841 aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa
     3901 agcaagtaaa acctctacaa atgtggtaaa atcgataagg atccgtcgac cgatgccctt
     3961 gagagccttc aacccagtca gctccttccg gtgggcgcgg ggcatgacta tcgtcgccgc
     4021 acttatgact gtcttcttta tcatgcaact cgtaggacag gtgccggcag cgctcttccg
     4081 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     4141 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     4201 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     4261 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     4321 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     4381 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     4441 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     4501 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     4561 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     4621 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     4681 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     4741 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     4801 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     4861 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     4921 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     4981 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     5041 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     5101 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     5161 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     5221 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     5281 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     5341 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     5401 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     5461 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     5521 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     5581 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     5641 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     5701 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     5761 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     5821 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     5881 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     5941 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     6001 cacctgacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     6061 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     6121 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     6181 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     6241 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     6301 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     6361 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     6421 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttgccattcg ccattcaggc
     6481 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagcccaagc
     6541 taccatgata agtaagtaat attaaggtac gggaggtact tggagcggcc gc
//
