LOCUS       sequence-138903-        6347 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For generating mRNA for zebrafish injection of cytoplasmic 
            fluorogen-activating protein (FAP).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6347)
  AUTHORS   He J, Wang Y, Missinato MA, Onuoha E, Perkins LA, Watkins SC, St 
            Croix CM, Tsang M, Bruchez MP
  TITLE     A genetically targetable near-infrared photosensitizer.
  JOURNAL   Nat Methods. 2016 Mar;13(3):263-268. doi: 10.1038/nmeth.3735. Epub 
            2016 Jan 25.
  PUBMED    26808669
REFERENCE   2  (bases 1 to 6347)
  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"; doi: 
            "10.1038/nmeth.3735"; journalName: "Nat Methods"; date: "2016-03"; 
            volume: "13"; issue: "3"; pages: "263-268"
COMMENT     Sequence Label: pCS2+ MBIC5-mCer3
FEATURES             Location/Qualifiers
     source          1..6347
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        638..656
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     681..697
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     682..698
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             2387..3106
                     /codon_start=1
                     /product="fluorogen-activating protein composed of a dimer 
                     of the L5-MG single-chain antibody fragment (Szent-Gyorgyi 
                     et al., 2008)"
                     /label=FAP-alpha-2
                     /note="binds malachite green derivatives to produce red 
                     fluorescence"
                     /translation="QAVVTQEPSVTVSPGGTVILTCGSGTGAVTSGHYANWFQQKPGQA
                     PRALIFDTDKKYSWTPGRFSGSLLGAKAALTISDAQPEDEAEYYCSLSDVDGYLFGGGT
                     QLTVLSGGGGSGGGGSGGGGSGGGGSQAVVTQEPSVTVSPGGTVILTCGSGTGAVTSGH
                     YANWFQQKPGQAPRALIFDTDKKYSWTPGRFSGSLLGAKAALTISDAQPEDEAEYYCSL
                     SDVDGYLFGGGTQLTVLS"
     CDS             join(3164..3166,3167..3169,3170..3883)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAIHGNVYITADKQKNGIK
                     ANFGLNCNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3209..3230)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3470..3489)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3817..3838
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3921..4055
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3945..3964)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3999..4018
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        complement(4158..4176)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4198..4214)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4198..4214)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4211..4233)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4222..4238
                     /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(4246..4252,4253..4270,4271..4276))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4291..4312
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4429..4446)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4600..5188)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4680..4699)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5359..6150,6151..6219))
                     /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     5982..6001
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6220..6324)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgaaa ttaccctgtt atccctataa tacgactcac tatagggcga
      661 attgggtacc gggccccccc tcgaggtcga cggtatcgat aagcttaaat cagttgtgtt
      721 aaataagaga cattcaaaat aaatgtaaat gagctctcca aatcagcaga cttaacattc
      781 tttaaaatga ttgattcaat agtgataaaa atcaggcata gccagttgta actttagata
      841 aattacagaa aatgtcaaat acagagaacc gattcttttt tatgatacat ccaagcacac
      901 atttaacaca atccaggcaa accccgaatt tcacagtcac aagcactgtt tgtacaagag
      961 ctttgcctaa ggacacacag tctctataag tccaggtcgt tggtttcact cttattttaa
     1021 acatgtgaca tttttcctgc catcctgtct taggctgctg tttgcttcat tccatgtcac
     1081 attaatttcc tcagtagcac cttttacaca cacagccaat cttttccaga aaattcaatt
     1141 gctttgaaaa gataatgtgt gaacaaatcc atttagaaaa ggaaaattaa gaatttgtaa
     1201 aatcatctgt aaattgttgg cattcttctg tatatgaaca tcacatcatt tacaggtaaa
     1261 ggtctggtca ttaattatat gacaatttac tggtattatt ttgtgaaagg ggctattttc
     1321 aatgcattca tccatccttt tcatccctca aatctctcat tcacgtcccc ctccccatct
     1381 gcacacttta tctcattttc caccctgctg gaatctgagc acttgtgcag ttatcagggc
     1441 tcctgtattt aggaggctct gggtgtccat gtaggggacg aacagaaaca ctgcagacct
     1501 ttatagaaga acaaatgata agagtcctca tacataaaga ctccattaga aacgtcagtg
     1561 acccaggagc ccagaccaac agcaaagcag acagtgaaca tggtgagtag acaaagctat
     1621 acttttttgg ttttgaatat aatattaatg tgaaaataaa aagcgtctat atgaagttaa
     1681 actaatcatt aaatagtgtt tgtttgttga tattaaatat tagaagcatc attttatgca
     1741 tttgtatgtt gtgatactat cttttaatga gtatttttac ttaggtactt ttttctgtac
     1801 actgtaaaac gcaaaaaagt taaggtaact caaatcattt gagaaaaccg attgtaacaa
     1861 gccatctgaa gttcaaaaac taatctaaat gagtactgtg aacttaatct atttgagtaa
     1921 aagaagcaat ttgagcacag taaaacccag taaatgaaga gaactcaacc aactgagtac
     1981 tgtaaaaccc aataagatga ggcagctcac accgtttgag gaaaccgatt gctacaaacc
     2041 atttgagtta aaaaaaaaga atcaatatga gtactgtgaa cttactccat ttaagttgaa
     2101 gtaatgaggt aattaattaa ctcattaccc tcaacactga gttcaaaact cttttcaaat
     2161 gagtagaatt aattttcagc caattttgag ttaactatac tcatttcatt tgataaagtt
     2221 gactattggg ttttacagtg tataaatgct ataaattgag tttccaaata aggcttctct
     2281 ttttttttac tcacaggcta gtaaaaaagc cgcggccaag agggggaaaa ctgctcaaag
     2341 aggctcttcc aatgtcttct ccatggaatt cggcctttct ggcctgcagg ccgtcgttac
     2401 ccaagaacct agtgttaccg ttagcccagg tggtactgtt atacttactt gtggaagtgg
     2461 tacgggtgcc gtcacatctg gtcattatgc aaattggttt caacaaaaac caggacaagc
     2521 tccaagagct ttgatttttg atactgataa gaagtattct tggaccccag gtagattttc
     2581 tggatctttg ctgggagcaa aggcagcttt gacaatatca gatgctcagc ctgaggacga
     2641 agccgagtat tactgttctc ttagcgacgt ggatggctac ttgtttggcg gtggaacaca
     2701 actgacggtt ctgtccggtg gtggcggctc tggtggcggt ggcagcggcg gtggtggttc
     2761 cggaggcggc ggttctcagg ctgtggtgac tcaggagccg tcagtgactg tgtccccagg
     2821 agggacagtc attctcactt gtggctccgg cactggagct gtcaccagtg gtcattatgc
     2881 caactggttc cagcagaagc ctggccaagc ccccagggca cttatatttg acaccgacaa
     2941 gaagtattcc tggacccctg gccgattctc aggctccctc cttggggcca aggctgccct
     3001 gaccatctcg gatgcgcagc ctgaagatga ggctgagtat tactgttcgc tctccgacgt
     3061 tgacggttat ctgttcggag gaggcaccca gctgaccgtc ctctccgcgg ccggtctggc
     3121 ccatatggga ggcggtgggt ctggtggcgg tggcagcacc ggtatggtga gcaagggcga
     3181 ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca
     3241 caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa
     3301 gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgag
     3361 ctggggcgtg cagtgcttcg cccgctaccc cgaccacatg aagcagcacg acttcttcaa
     3421 gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa
     3481 ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct
     3541 gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacgc
     3601 catccacggc aacgtctata tcaccgccga caagcagaag aacggcatca aggccaactt
     3661 cggcctcaac tgcaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa
     3721 cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc
     3781 caagctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac
     3841 cgccgccggg atcactctcg gcatggacga gctgtacaag taactcgagc ctctagaact
     3901 atagtgagtc gtattacgta gatccagaca tgataagata cattgatgag tttggacaaa
     3961 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     4021 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     4081 tgtttcaggt tcagggggag gtgtgggagg ttttttaatt cgcggccgcc accgcggtgg
     4141 agctccagct tttgttccct ttagtgaggg ttaattaccc tgttatccct aatcatggtc
     4201 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     4261 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     4321 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     4381 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     4441 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     4501 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     4561 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     4621 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     4681 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     4741 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     4801 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     4861 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     4921 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     4981 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     5041 gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     5101 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     5161 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     5221 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     5281 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     5341 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     5401 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     5461 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     5521 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     5581 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     5641 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     5701 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     5761 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     5821 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     5881 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     5941 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     6001 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     6061 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     6121 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     6181 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     6241 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     6301 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccac
//
