LOCUS       sequence-227832-        8360 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CAG:mcherry:P2A:HA_NLGN.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8360)
  TITLE     SYNPLA plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8360)
  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            "Unpublished"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: jk96
FEATURES             Location/Qualifiers
     source          1..8360
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   259..388
                     /label=AAV2 ITR
     promoter        701..978
                     /label=chicken beta-actin promoter
     primer_bind     1258..1280
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1342..1361
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1394..1403
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1400..2107
                     /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(1545..1563)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1813..1832
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2120..2176
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2318..2344
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     regulatory      2806..2815
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    4754..5342
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4807..4827)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5225..5236)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    5385..5506
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5396..5415)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5450..5469
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   5668..5797
                     /label=AAV2 ITR
     primer_bind     complement(5816..5833)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5816..5832)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5825..5847)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5974..6429
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6061..6080)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6271..6292
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6455..6559
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6560..6628,6629..7420)
                     /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(6778..6797)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7591..8179
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8080..8099
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccagattta
      241 attaaggcct taattaggct gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc
      301 gggcgtcggg cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga
      361 gtggccaact ccatcactag gggttccttg tagttaatga ttaacccgcc atgctactta
      421 tctacgtagc catgctctag gaagatcgta ccattgacgt caataatgac gtatgttccc
      481 atagtaacgc caatagggac tttccattga cgtcaatggg tggactattt acggtaaact
      541 gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat
      601 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact
      661 tggcagtaca tctacgtatt agtcatcgct attaccatgg tcgaggtgag ccccacgttc
      721 tgcttcactc tccccatctc ccccccctcc ccacccccaa ttttgtattt atttattttt
      781 taattatttt gtgcagcgat gggggcgggg gggggggggg ggcgcgcgcc aggcggggcg
      841 gggcggggcg aggggcgggg cggggcgagg cggagaggtg cggcggcagc caatcagagc
      901 ggcgcgctcc gaaagtttcc ttttatggcg aggcggcggc ggcggcggcc ctataaaaag
      961 cgaagcgcgc ggcgggcggg agtcgctgcg acgctgcctt cgccccgtgc cccgctccgc
     1021 cgccgcctcg cgccgcccgc cccggctctg actgaccgcg ttactcccac aggtgagcgg
     1081 gcgggacggc ccttctcctc cgggctgtaa ttagcgcttg gtttaatgac ggcttgtttc
     1141 ttttctgtgg ctgcgtgaaa gccttgaggg gctccgggag ggccctttgt gcggggggag
     1201 cggctcgggg ctgtccgcgg ggggacggct gccttcgggg gggacggggc agggcggggt
     1261 tcggcttctg gcgtgtgacc ggcggctcta gagcctctgc taaccatgtt catgccttct
     1321 tctttttcct acagctcctg ggcaacgtgc tggttattgt gctgtctcat cattttggca
     1381 aagaattgga tctgccacca tggtgagcaa gggcgaggag gataacatgg ccatcatcaa
     1441 ggagttcatg cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat
     1501 cgagggcgag ggcgagggcc gcccctacga gggcacccag accgccaagc tgaaggtgac
     1561 caagggtggc cccctgccct tcgcctggga catcctgtcc cctcagttca tgtacggctc
     1621 caaggcctac gtgaagcacc ccgccgacat ccccgactac ttgaagctgt ccttccccga
     1681 gggcttcaag tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca
     1741 ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt
     1801 cccctccgac ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct cctccgagcg
     1861 gatgtacccc gaggacggcg ccctgaaggg cgagatcaag cagaggctga agctgaagga
     1921 cggcggccac tacgacgctg aggtcaagac cacctacaag gccaagaagc ccgtgcagct
     1981 gcccggcgcc tacaacgtca acatcaagtt ggacatcacc tcccacaacg aggactacac
     2041 catcgtggaa cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct
     2101 gtacaagtct ggaagcggag ctactaactt cagcctgctg aagcaggctg gagacgtgga
     2161 ggagaaccct ggacctatgg cacttcccag atgcatgtgg ccaaattatg tttggagagc
     2221 tatgatggca tgtgtggtcc acaggggatc cggtgcccca ttgactctct gcttgttggg
     2281 atgtttgcta cagacttttc acgtactctc tcaaaagtat ccatacgacg ttccggacta
     2341 cgcagtcgac ttggatgatg tagacccatt ggttactact aactttggca agattagggg
     2401 aattaagaaa gaactcaata atgaaatttt gggtcctgtc attcagtttc ttggggttcc
     2461 atatgccgct ccaccaacag gagaacatcg tttccagcct ccagaaccac catctccctg
     2521 gtctgacatc cggaacgcca ctcagtttgc tcctgtatgt ccccagaata tcattgatgg
     2581 cagattgcct gaagttatgc ttcctgtgtg gttcactaat aacttggatg tggtttcatc
     2641 atacgtccaa gaccagagtg aagactgtct atacttaaac atctatgtcc caactgaaga
     2701 tggtcccctt acaaagaaac acacagatga tttaggtgat aatgacggtg ctgaagatga
     2761 agatattcgg gacagtgggg gtcccaaacc agtgatggtg tacatccatg gcggctctta
     2821 catggaaggt actggaaatc tgtatgatgg gagtgtcttg gcaagctatg gcaatgtgat
     2881 cgtcatcaca gtcaactatc ggcttggggt acttggcttc ttgagcacag gggatcaggc
     2941 tgccaaagga aactacgggc tccttgacct catccaggcc ctaagatgga ccagcgagaa
     3001 cattgggttc tttggtggtg accccttgcg aatcactgtg tttggatcag gcgctggggg
     3061 ttcatgtgtc aacctgctga ctttatccca ttattctgaa ggtaaccgtt ggagcaattc
     3121 aaccaaagga ctttttcaac gagcaatagc tcagagtgga acagcccttt ccagctgggc
     3181 tgttagtttc cagcctgcaa aatacgctag aattctggcc acaaaagttg gctgcaatgt
     3241 ttcagataca gtagagttag tagaatgcct gcagaagaag ccttacaaag aacttgttga
     3301 tcaagatgtt caaccagccc gataccacat agcctttgga cctgtgatcg atggtgatgt
     3361 aataccagat gaccctcaga tactgatgga acaaggagag ttcctcaact atgatataat
     3421 gttgggagtt aaccaagggg aagggttgaa gtttgtcgaa aatatagtag atagtgatga
     3481 tggtgtatca gccagtgatt tcgactttgc tgtttctaat tttgttgata atttatatgg
     3541 atatcctgaa ggcaaagatg ttttgagaga aaccattaaa ttcatgtata ctgactgggc
     3601 tgatcgccat aatcctgaaa ctagaaggaa gacattgttg gctttgttta cggaccatca
     3661 atgggtagca cctgctgtgg ccacagcaga ccttcactcg aactttggct cacctacata
     3721 cttctatgcc ttttatcatc attgccaaac agatcaagtt ccagcttggg ctgatgcagc
     3781 tcatggggat gaggttccct atgtgttggg aatccccatg attggcccta cagagttatt
     3841 tccttgcaat ttctccaaga atgatgtgat gttgagtgca gtagtaatga catactggac
     3901 gaattttgct aaaactggtg acccaaatca accagttcct caagacacaa aattcatcca
     3961 taccaaaccc aaccgctttg aagaagtagc atggaccaga tattcccaga aagaccagct
     4021 ttatctccat attggattaa aaccgagagt taaagagcat tacagagcca ataaggtaaa
     4081 tctctggctg gagctggtac ctcatctgca taatctcaat gacatttctc agtatacctc
     4141 gacaacaact aaagtgccat ccacggacat cactctcaga cctacaagga agaattccac
     4201 accagtcaca tcagcctttc ccactgccaa acaggatgat cccaagcaac aaccaagccc
     4261 cttctcggtg gatcagaggg actactccac agagctaagt gtcactatcg cagtgggggc
     4321 ctctctgctg tttctcaaca tcttggcttt tgcagccctg tactacaaga aggataagag
     4381 gagacatgat gtccaccgga ggtgcagccc tcagcgcacg accaccaacg acctaaccca
     4441 tgctccagaa gaggaaatta tgtctctcca aatgaagcac actgacttgg atcacgagtg
     4501 tgagtccatc catccacatg aggtggttct tcggaccgcc tgtcccccag attatactct
     4561 agctatgagg aggtcacctg atgatattcc actaatgaca cctaacacca tcacaatgat
     4621 tcccaacact ataccaggga ttcagccctt acatacattc aacacattta ctggaggaca
     4681 gaataataca ctgccccatc cccacccaca cccccattca cattcaacaa ccagggtata
     4741 gaagcttatc gataatcaac ctctggatta caaaatttgt gaaagattga ctggtattct
     4801 taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc
     4861 tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct
     4921 ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga
     4981 cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc
     5041 tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac
     5101 aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaaat catcgtcctt
     5161 tccttggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt
     5221 cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc
     5281 tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc
     5341 gcatcgatac cgtcgacccg ggcggccgct tcgagcagac atgataagat acattgatga
     5401 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga
     5461 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg
     5521 cattcatttt atgtttcagg ttcaggggga gatgtgggag gttttttaaa gcaagtaaaa
     5581 cctctacaaa tgtggtaaaa tcgataagga tcttcctaga gcatggctac gtagataagt
     5641 agcatggcgg gttaatcatt aactacaagg aacccctagt gatggagttg gccactccct
     5701 ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct
     5761 ttgcccgggc ggcctcagtg agcgagcgag cgcgcagcct taattaacct aattcactgg
     5821 ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg
     5881 cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt
     5941 cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg
     6001 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     6061 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     6121 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     6181 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     6241 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     6301 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     6361 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc
     6421 ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     6481 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     6541 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     6601 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     6661 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     6721 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     6781 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     6841 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     6901 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     6961 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     7021 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     7081 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     7141 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     7201 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     7261 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     7321 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     7381 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     7441 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     7501 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     7561 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     7621 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     7681 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     7741 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     7801 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     7861 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     7921 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     7981 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     8041 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     8101 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     8161 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     8221 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     8281 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     8341 agtgagcgag gaagcggaag
//
