LOCUS       sequence-176064-        7715 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  encodes human/murine miR-146a and ZsGreen1 .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7715)
  TITLE     pAPM-mmu-miR146a
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7715)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pALPS-ZsGreen-miR-146a
FEATURES             Location/Qualifiers
     source          1..7715
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     207..227
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     primer_bind     401..420
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        495..599
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(600..668,669..1460)
                     /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(818..837)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1631..2219
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2120..2139
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     LTR             2290..2470
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    2988..3576
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             3094..3105
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     3503..3523
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4019..4714)
                     /codon_start=1
                     /product="Zoanthus green fluorescent protein"
                     /label=ZsGreen1
                     /note="mammalian codon-optimized"
                     /translation="MAQSKHGLTKEMTMKYRMEGCVDGHKFVITGEGIGYPFKGKQAIN
                     LCVVEGGPLPFAEDILSAAFMYGNRVFTEYPQDIVDYFKNSCPAGYTWDRSFLFEDGAV
                     CICNADITVSVEENCMYHESKFYGVNFPADGPVMKKMTDNWEPSCEKIIPVPKQGILKG
                     DVSMYLLLKDGGRLRCQFDTVYKAKSVPRKMPDWHFIQHKLTREDRSDAKNQKWHLTEH
                     AIASGSALP"
     regulatory      4711..4720
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(4729..5136)
                     /label=SFFV promoter
                     /note="spleen focus-forming virus long terminal repeat 
                     (LTR) promoter"
     misc_feature    5273..5390
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     CDS             complement(5688..5732)
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    5917..6150
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    6647..6772
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             6819..6999
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(7017..7215)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(7042..7062)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        7217..7596
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        complement(join(7608..7715,1..222))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(7622..7715,1..42)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(7676..7695)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 gggcggagaa tgggcggaac tgggcggagt taggggcggg atgggcggag ttaggggcgg
       61 gactatggtt gctgactaat tgagatgcat gctttgcata cttctgcctg ctggggagcc
      121 tggggacttt ccacacctgg ttgctgacta attgagatgc atgctttgca tacttctgcc
      181 tgctggggag cctggggact ttccacaccc taactgacac acattccaca gcgcgtatat
      241 ctggcccgta catcgcgaag cagcgcaaaa cgcctaaccc taagcagatt cttcatgcaa
      301 ttgtcggtca agccttgcct tgttgtagct taaattttgc tcgcgcacta ctcagcgacc
      361 tccaacacac aagcagggag cagatactgg cttaactatg cggcatcaga gcagattgta
      421 ctgagagtgc accatagggg atcgggagat ctcccgatcc gtcgacgtca ggtggcactt
      481 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
      541 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
      601 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
      661 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
      721 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
      781 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
      841 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
      901 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
      961 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     1021 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     1081 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     1141 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     1201 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     1261 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     1321 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     1381 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     1441 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     1501 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     1561 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     1621 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     1681 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     1741 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     1801 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     1861 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     1921 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     1981 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     2041 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     2101 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     2161 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     2221 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatga
     2281 aacgggccct gctagagatt ttccacactg actaaaaggg tctgagggat ctctagttac
     2341 cagagtcaca caacagacgg gcacacacta cttgaagcac tcaaggcaag ctttattgag
     2401 gcttaagcag tgggttccct agttagccag agagctccca ggctcagatc tggtctaacc
     2461 agagagaccc agtacagtcc ggatgcagct ctcgggccat gtgatgaaat gctaggcggc
     2521 tgtcaaacct ccactctaat acttctctct ccgggtcatc catcccatgc aggctcacag
     2581 ggtgtaacaa gcgggtgttc tctccttcat tggcttcttc taccttctct tgctcaactg
     2641 gtactagctt gtagcaccat ccaaaggtca gtggatatct gatccctggc cctggtgtgt
     2701 agttctgcca atcagggaag tagccttgtg tgtggtagat ccacagatca aggatatctt
     2761 gtcttcgttg ggagtgaatt agcccttcca gtcccccctt ttcttttaaa aagtggctaa
     2821 gatctacagc tgccttgtaa gtcattggtc ttaaaggtac ctgaggtgtg actggaaaac
     2881 ccacctcctc ctcctcttgt gcttctagcc aggcacaatc agcattggta gctgctgtat
     2941 tgctacttgt gattgctcca tgtttttcta ggtctcgacg cgattaagcg gggaggcggc
     3001 ccaaagggag atccgactcg tctgagggcg aaggcgaaga cgcggaagag gccgcagagc
     3061 cggcagcagg ccgcgggaag gaaggtccgc tggattgagg gccgaaggga cgtagcagaa
     3121 ggacgtcccg cgcagaatcc aggtggcaac acaggcgagc agccatggaa aggacgtcag
     3181 cttccccgac aacaccacgg aattgtcagt gcccaacagc cgagcccctg tccagcagcg
     3241 ggcaaggcag gcggcgatga gttccgccgt ggcaataggg agggggaaag cgaaagtccc
     3301 ggaaaggagc tgacaggtgg tggcaatgcc ccaaccagtg ggggttgcgt cagcaaacac
     3361 agtgcacacc acgccacgtt gcctgacaac gggccacaac tcctcataaa gagacagcaa
     3421 ccaggattta tacaaggagg agaaaatgaa agccatacgg gaagcaatag catgatacaa
     3481 aggcattaaa gcagcgtatc cacatagcgt aaaaggagca acatagttaa gaataccagt
     3541 caatctttca caaattttgt aatccagagg ttgattagcg atcgcggccc caggggccac
     3601 cggtatcgat acgcgtcaca tcaatccaac atgactccct ccggtgccag cagttccacg
     3661 cttcacccca ctctctccac tcttcaagtt ccttggcaga ctatgaaggc cttcagagtt
     3721 tgttccaacc agtgttcttc aggtccgtga tgacagagct atcccagctg aagaactgaa
     3781 tttcacaggt ctgacattga tataacccat ggaattcagt tctcagagct ggggatacac
     3841 acagactctc cagatgaggc cttcctccca ggccagatcc tgccagccct gtaaaaacac
     3901 aagagaaatc tctgataaat ccaaaagcac ggcctctgca gactggacac tcttgatgca
     3961 taaggatctg ggacacaaga taggcaagtt cataagcaat cattaaacat gcggccgctt
     4021 agggcaaggc ggagccggag gcgatggcgt gctcggtcag gtgccacttc tggttcttgg
     4081 cgtcgctgcg gtcctcgcgg gtcagcttgt gctggatgaa gtgccagtcg ggcatcttgc
     4141 ggggcacgga cttggccttg tacacggtgt cgaactggca gcgcaagcgg ccaccgtcct
     4201 tcagcagcag gtacatgctc acgtcgccct tcaagatgcc ctgcttgggc acggggatga
     4261 tcttctcgca ggagggctcc cagttgtcgg tcatcttctt catcacgggg ccgtcggcgg
     4321 ggaagttcac gccgtagaac ttggactcgt ggtacatgca gttctcctcc acgctcacgg
     4381 tgatgtcggc gttgcagatg cacacggcgc cgtcctcgaa caggaaggag cggtcccagg
     4441 tgtagccggc ggggcaggag ttcttgaagt agtcgacgat gtcctggggg tactcggtga
     4501 acacgcggtt gccgtacatg aaggcggcgg acaagatgtc ctcggcgaag ggcaaggggc
     4561 cgccctccac cacgcacagg ttgatggcct gcttgccctt gaaggggtag ccgatgccct
     4621 cgccggtgat cacgaacttg tggccgtcca cgcagccctc catgcggtac ttcatggtca
     4681 tctccttggt caggccgtgc ttggactggg ccatggtggc tctagacccc cgggcgactc
     4741 agtctgtcgg aggactggcg cgccgagtga ggggttgtga gctcttttat agagctcggg
     4801 aagcagaagc gcgcgaacag aagcgagaag caggctgatt ggttaattca aataaggcgc
     4861 agggtcattt caggtccttg ggggagcctg gaaacatctg atgggtctta agaaactgct
     4921 gagggttggg ccatatctgg ggaccatctg ttcttggcct cgggccgggg ccgaaactgc
     4981 ggtgaccatc tgttcttggc cccgggccgg ggccgaaact gctcaccgca gatatcctgt
     5041 ttggcccaac gttagctatt ttcatgtacc cgcccttgat ctgaacttct ctattcttgg
     5101 tttggtattt ttccatgcct tgcaaaatgg cgttactgca gctagcttgc caaacctaca
     5161 ggtggggtct ttcattcccc cctttttctg gagactaaat aaaatctttt attttatcgg
     5221 ggctgcagtt aattaaccaa actggatctc tgctgtccct gtaataaacc cgaaaatttt
     5281 gaatttttgt aatttgtttt tgtaattctt tagtttgtat gtctgttgct attatgtcta
     5341 ctattctttc ccctgcactg taccccccaa tccccccttt tcttttaaaa ttgtggatga
     5401 atactgccat ttgtctgcag aattggcgca cgcagtgccg atccgttcac taatcgaatg
     5461 gatctgtctc tgtctctctc tccaccttct tcttctattc cttcgggcct gtcgggtccc
     5521 ctcggggttg ggaggtgggt ctgaaacgat aatggtgaat atccctgcct aactctattc
     5581 actatagaaa gtacagcaaa aactattctt aaacctacca agcctcctac tatcattatg
     5641 aataatttta tataccacag ccaatttgtt atgttaaacc aattccacaa acttgcccat
     5701 ttatctaatt ccaataattc ttgttcattc ttttcttgct ggttttgcga ttcttcaatt
     5761 aaggagtgta ttaagcttgt gtaattgtta atttctctgt cccactccat ccaggtcgtg
     5821 tgattccaaa tctgttccag agatttatta ctccaactag cattccaagg cacagcagtg
     5881 gtgcaaatga gttttccaga gcaaccccaa atccccagga gctgttgatc ctttaggtat
     5941 ctttccacag ccaggattct tgcctggagc tgcttgatgc cccagactgt gagttgcaac
     6001 agatgctgtt gcgcctcaat agccctcagc aaattgttct gctgctgcac tataccagac
     6061 aataattgtc tggcctgtac cgtcagcgtc attgacgctg cgcccatagt gcttcctgct
     6121 gctcccaaga acccaaggaa caaagctcct attcccactg ctcttttttc tctctgcacc
     6181 actcttctct ttgccttggt gggtgctact cctaatggtt caatttttac tactttatat
     6241 ttatataatt cacttctcca attgtccctc atatctcctc ctccaggtct gaagatcagc
     6301 ggccggccgc ttgctgtgcg gtggtcttac ttttgttttg ctcttcctct atcttgtcta
     6361 aagcttcctt ggtgtctttt atctctatcc tttgatgcac acaatagagg gttgctactg
     6421 tattatataa tgatctaagt tcttctgatc ctgtctgaag ggatggttgt agctgtccca
     6481 gtatttgtct acagccttct gatgtttcta acaggccagg attaactgcg aatcgttcta
     6541 gctccctgct tgcccatact atatgtttta atttatattt tttctttccc cctggcctta
     6601 accgaatttt ttcccatcgc gatctaattc tcccccgctt aatactgacg ctctcgcacc
     6661 catctctctc cttctagcct ccgctagtca aaatttttgg cgtactcacc agtcgccgcc
     6721 cctcgcctct tgccgtgcgc gcttcagcaa gccgagtcct gcgtcgagag agctcctctg
     6781 gtttcccttt cgctttcaag tccctgttcg ggcgccactg ctagagattt tccacactga
     6841 ctaaaagggt ctgagggatc tctagttacc agagtcacac aacagacggg cacacactac
     6901 ttgaagcact caaggcaagc tttattgagg cttaagcagt gggttcccta gttagccaga
     6961 gagctcccag gctcagatct ggtctaacca gagagaccca gtacaggcaa aacgcgctgc
     7021 ttatatagac ctcccaccgt acacgcctac cgcccatttg cgtcaatggg gcggagttgt
     7081 tacgacattt tggaaagtcc cgttgatttt ggtgccaaaa caaactccca ttgacgtcaa
     7141 tggggtggag acttggaaat ccccgtgagt caaaccgcta tccacgccca ttgatgtact
     7201 gccaaaaccg catcaccatg gtaatagcga tgactaatac gtagatgtac tgccaagtag
     7261 gaaagtccca taaggtcatg tactgggcat aatgccaggc gggccattta ccgtcattga
     7321 cgtcaatagg gggcgtactt ggcatatgat acacttgatg tactgccaag tgggcagttt
     7381 accgtaaata ctccacccat tgacgtcaat ggaaagtccc tattggcgtt actatgggaa
     7441 catacgtcat tattgacgtc aatgggcggg ggtcgttggg cggtcagcca ggcgggccat
     7501 ttaccgtaag ttatgtaacg cggaactcca tatatgggct atgaactaat gaccccgtaa
     7561 ttgattacta ttaataacta gtcaataatc aatgtcaacg cgagcttttt gcaaaagcct
     7621 aggcctccaa aaaagcctcc tcactacttc tggaatagct cagaggcaga ggcggcctcg
     7681 gcctctgcat aaataaaaaa aattagtcag ccatg
//
