LOCUS       sequence-256379-        7991 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Piggybac vector for constitutive expression of iRFP670-NLS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7991)
  TITLE     Madison Lab plasmids
REFERENCE   2  (bases 1 to 7991)
  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: BII-ChPtW-iRFP670
FEATURES             Location/Qualifiers
     source          1..7991
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   5..39
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(308..324)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(308..324)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             1875..1928
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             1935..1961
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             1935..1961
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1968..2912
                     /codon_start=1
                     /product="bright monomeric near-infrared fluorescent 
                     protein engineered from a bacterial phytochrome, with an 
                     emission maximum of 670 nm (Shcherbakova et al., 2016)"
                     /label=miRFP670
                     /label=ORF frame 1
                     /translation="MVAGHASGSPAFGTASHSNCEHEEIHLAGSIQPHGALLVVSEHDH
                     RVIQASANAAEFLNLGSVLGVPLAEIDGDLLIKILPHLDPTAEGMPVAVRCRIGNPSTE
                     YCGLMHRPPEGGLIIELERAGPSIDLSGTLAPALERIRTAGSLRALCDDTVLLFQQCTG
                     YDRVMVYRFDEQGHGLVFSECHVPGLESYFGNRYPSSTVPQMARQLYVRQRVRVLVDVT
                     YQPVPLEPRLSPLTGRDLDMSGCFLRSMSPCHLQFLKDMGVRATLAVSLVVGGKLWGLV
                     VCHHYLPRFIRFELRAICKRLAERIATRITALES"
     CDS             2916..2936
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     misc_feature    2968..3556
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3021..3041)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3439..3450)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3636..3770
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3660..3679)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3714..3733
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4548..4610
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(4740..4757)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4740..4756)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4749..4771)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4898..5353
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4985..5004)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5195..5216
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5379..5483
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5484..5552,5553..6344)
                     /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(5702..5721)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6515..7103
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7004..7023
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7257..7274
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7391..7412
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7427..7432,7433..7450,7451..7457)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7465..7481
                     /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     7470..7492
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7489..7505
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7489..7505
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttaaccctag aaagatagtc tgcgtaaaat tgacgcatgc attcttgaaa tattgctctc
       61 tctttctaaa tagcgcgaat ccgtcgctgt gcatttagga catctcagtc gccgcttgga
      121 gctcccgtga ggcgtgcttg tcaatgcggt aagtgtcact gattttgaac tataacgacc
      181 gcgtgagtca aaatgacgca tgattatctt ttacgtgact tttaagattt aactcatacg
      241 ataattatat tgttatttca tgttctactt acgtgataac ttattatata tatattttct
      301 tgttatagat ccactagttc tagagggaca gccccccccc aaagccccca gggatgtaat
      361 tacgtccctc ccccgctagg gggcagcagc gagccgcccg gggctccgct ccggtccggc
      421 gctccccccg catccccgag ccggcagcgt gcggggacag cccgggcacg gggaaggtgg
      481 cacgggatcg ctttcctctg aacgcttctc gctgctcttt gagcctgcag acacctgggg
      541 ggatacgggg aaaaggcctc caaggcctgc aggagtcaat gggaaaaacc cattggagcc
      601 aagtacactg actcaatagg gactttccat tgggttttgc ccagtacata aggtcaatag
      661 ggggtgagtc aacaggaaag tcccattgga gccaagtaca ttgagtcaat agggactttc
      721 caatgggttt tgcccagtac ataaggtcaa tgggaggtaa gccaatgggt ttttcccatt
      781 actgacatgt atactgagtc attagggact ttccaatggg ttttgcccag tacataaggt
      841 caataggggt gaatcaacag gaaagtccca ttggagccaa gtacactgag tcaataggga
      901 ctttccattg ggttttgccc agtacaaaag gtcaataggg ggtgagtcaa tgggtttttc
      961 ccattattgg cacatacata aggtcaatag gggtgactag tggagaagag catgcttgag
     1021 ggctgagtgc ccctcagtgg gcagagagca catggcccac agtccctgag aagttggggg
     1081 gaggggtggg caattgaact ggtgcctaga gaaggtgggg cttgggtaaa ctgggaaagt
     1141 gatgtggtgt actggctcca cctttttccc cagggtgggg gagaaccata tataagtgca
     1201 gtagtctctg tgaacattcg gtttagtgaa cctgcaggtc ggccgccacg accggtgcca
     1261 ccatgactga atacaaacca actgttcgcc tggcaactcg tgatgatgtt ccacgtgcag
     1321 ttcgcaccct ggctgctgca tttgctgact accctgcaac ccgtcacact gtggacccag
     1381 accgccacat tgaacgtgtg actgaactgc aggagctgtt cctgacccgt gtgggcctgg
     1441 acattggcaa agtgtgggtg gcagatgatg gtgctgctgt ggcagtgtgg accacccctg
     1501 aatctgttga agctggtgca gtgtttgctg agattggccc acgcatggca gaactgtctg
     1561 gcagccgcct ggcagcacaa cagcagatgg aaggtctgct ggcaccacac cgcccaaaag
     1621 aacctgcttg gttcctggca actgtgggtg tgagccctga ccaccagggt aagggcctgg
     1681 gctctgcagt ggtgctgcct ggtgtggaag cagctgaacg tgcaggtgtg cctgctttcc
     1741 tggagacctc agctccacgc aacctgcctt tctatgaacg cctgggcttc actgtgactg
     1801 ctgatgtgga agtgccagaa ggcccacgca cttggtgcat gactcgcaaa ccaggtgcta
     1861 gccttggctc cggcgagggc aggggaagtc ttctaacatg cggggacgtg gaggaaaatc
     1921 ccggcccact cggctacccc tacgacgtgc ccgactacgc cagcggcatg gtagcaggtc
     1981 atgcctctgg cagccccgca ttcgggaccg cctctcattc gaattgcgaa catgaagaga
     2041 tccacctcgc cggctcgatc cagccgcatg gcgcgcttct ggtcgtcagc gaacatgatc
     2101 atcgcgtcat ccaggccagc gccaacgccg cggaatttct gaatctcgga agcgtactcg
     2161 gcgttccgct cgccgagatc gacggcgatc tgttgatcaa gatcctgccg catctcgatc
     2221 ccaccgccga aggcatgccg gtcgcggtgc gctgccggat cggcaatccc tctacggagt
     2281 actgcggtct gatgcatcgg cctccggaag gcgggctgat catcgaactc gaacgtgccg
     2341 gcccgtcgat cgatctgtca ggcacgctgg cgccggcgct ggagcggatc cgcacggcgg
     2401 gttcactgcg cgcgctgtgc gatgacaccg tgctgctgtt tcagcagtgc accggctacg
     2461 accgggtgat ggtgtatcgt ttcgatgagc aaggccacgg cctggtattc tccgagtgcc
     2521 atgtgcctgg gctcgaatcc tatttcggca accgctatcc gtcgtcgact gtcccgcaga
     2581 tggcgcggca gctgtacgtg cggcagcgcg tccgcgtgct ggtcgacgtc acctatcagc
     2641 cggtgccgct ggagccgcgg ctgtcgccgc tgaccgggcg cgatctcgac atgtcgggct
     2701 gcttcctgcg ctcgatgtcg ccgtgccatc tgcagttcct gaaggacatg ggcgtgcgcg
     2761 ccaccctggc ggtgtcgctg gtggtcggcg gcaagctgtg gggcctggtt gtctgtcacc
     2821 attatctgcc gcgcttcatc cgtttcgagc tgcgggcgat ctgcaaacgg ctcgccgaaa
     2881 ggatcgcgac gcggatcacc gcgcttgaga gcgggcccaa gaagaagcgc aaggtgggcg
     2941 cctaaggcta gagttcgcgt ctggaacaat caacctctgg attacaaaat ttgtgaaaga
     3001 ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg
     3061 cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc
     3121 tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc
     3181 actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt
     3241 tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt
     3301 gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg
     3361 aagctgacgt cctttccatg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg
     3421 tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg
     3481 ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt
     3541 tgggccgcct ccccgcctgg aattaattct ggctaagcac ttcgtggccg tcgatcgttt
     3601 aaagggaggt agtgagtcga gatatcgata ctagtgatcc agacatgata agatacattg
     3661 atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt
     3721 gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca
     3781 attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt taaagcaagt
     3841 aaaacctcta caaatgtggt atggctgatt atgatcctct agagggacag ccccccccca
     3901 aagcccccag ggatgtaatt acgtccctcc cccgctaggg ggcagcagcg agccgcccgg
     3961 ggctccgctc cggtccggcg ctccccccgc atccccgagc cggcagcgtg cggggacagc
     4021 ccgggcacgg ggaaggtggc acgggatcgc tttcctctga acgcttctcg ctgctctttg
     4081 agcctgcaga cacctggggg gatacgggga aaagccatac caatggagct cacggggaca
     4141 gccccccccc aaagccccca gggatgtaat tacgtccctc ccccgctagg gggcagcagc
     4201 gagccgcccg gggctccgct ccggtccggc gctccccccg catccccgag ccggcagcgt
     4261 gcggggacag cccgggcacg gggaaggtgg cacgggatcg ctttcctctg aacgcttctc
     4321 gctgctcttt gagcctgcag acacctgggg ggatacgggg aaaaagaatt cgataaaagt
     4381 tttgttactt tatagaagaa attttgagtt tttgtttttt ttaataaata aataaacata
     4441 aataaattgt ttgttgaatt tattattagt atgtaagtgt aaatataata aaacttaata
     4501 tctattcaaa ttaataaata aacctcgata tacagaccga taaaacacat gcgtcaattt
     4561 tacacatgat tatctttaac gtacgtcaca atatgattat ctttctaggg ttaatctagc
     4621 tgcgtgttct gcagcgtgtc gagcatcttc atctgctcca tcacgctgta aaacacattt
     4681 gcaccgcgag tctgcccgtc ctccacgggt tcaaaaacgt gaatgaacga ggcgcgctca
     4741 ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc
     4801 cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc
     4861 ccttcccaac agttgcgcag cctgaatggc gaatgggacg cgccctgtag cggcgcatta
     4921 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     4981 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     5041 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     5101 aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     5161 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     5221 acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc
     5281 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     5341 acgcttacaa tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     5401 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     5461 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     5521 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     5581 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     5641 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     5701 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     5761 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     5821 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     5881 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     5941 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     6001 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
     6061 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     6121 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     6181 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     6241 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     6301 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     6361 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     6421 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     6481 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     6541 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     6601 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     6661 tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     6721 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     6781 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     6841 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     6901 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     6961 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     7021 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     7081 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     7141 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
     7201 gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg
     7261 agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt
     7321 ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc
     7381 gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc
     7441 ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct
     7501 atgaccatga ttacgccaag cgcgcccgcc gggtaactca cggggtatcc atgtccattt
     7561 ctgcggcatc cagccaggat acccgtcctc gctgacgtaa tatcccagcg ccgcaccgct
     7621 gtcattaatc tgcacaccgg cacggcagtt ccggctgtcg ccggtattgt tcgggttgct
     7681 gatgcgcttc gggctgacca tccggaactg tgtccggaaa agccgcgacg aactggtatc
     7741 ccaggtggcc tgaacgaaca gttcaccgtt aaaggcgtgc atggccacac cttcccgaat
     7801 catcatggta aacgtgcgtt ttcgctcaac gtcaatgcag cagcagtcat cctcggcaaa
     7861 ctctttccat gccgcttcaa cctcgcggga aaaggcacgg gcttcttcct ccccgatgcc
     7921 cagatagcgc cagcttgggc gatgactgag ccggaaaaaa gacccgacga tatgatcctg
     7981 atgcagctag a
//
