LOCUS       sequence-431538-        3397 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fragmid fragment: (2A Selection) TagRFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3397)
  AUTHORS   McGee AV, Liu YV, Griffith AL, Szegletes ZM, Wen B, Kraus C, Miller 
            NW, Steger RJ, Escude Velasco B, Bosch JA, Zirin JD, Viswanatha R, 
            Sontheimer EJ, Goodale A, Greene MA, Green TM, Doench JG
  TITLE     Modular vector assembly enables rapid assessment of emerging CRISPR 
            technologies.
  JOURNAL   Cell Genom. 2024 Mar 13;4(3):100519. doi: 
            10.1016/j.xgen.2024.100519.
  PUBMED    38484704
REFERENCE   2  (bases 1 to 3397)
  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: "Cell 
            Genom."; date: "2024-03-13"; volume: "4(3):100519. doi"; pages: " 
            10.1016/j.xgen.2024.100519"
COMMENT     Sequence Label: AA266
FEATURES             Location/Qualifiers
     source          1..3397
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             444..500
                     /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             513..1226
                     /codon_start=1
                     /product="monomeric derivative of red fluorescent protein 
                     from Entacmaea quadricolor (Merzlyak et al., 2007)"
                     /label=TagRFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTM
                     RIKVVEGGPLPFAFDILATSFMYGSRTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEANTEMLYPADGGLEGRSD
                     MALKLVGGGHLICNFKTTYRSKKPAKNLKMPGVYYVDHRLERIKEADKETYVEQHEVAV
                     ARYCDLPSKLGHKLN"
     primer_bind     complement(1307..1323)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1307..1323)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1320..1342)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1331..1347
                     /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(1355..1361,1362..1379,1380..1385))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1400..1421
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1538..1555)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1709..2297)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1789..1808)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2475..3284)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3198..3217
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3276..3295
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagagaatt cgagctcggt acctcgcgaa
      421 tacatctaga tgaagacgga agcgcaacaa acttctctct gctgaaacaa gccggagatg
      481 tcgaagagaa tcctggaccg gcaccgggat ccatggtgtc taagggcgaa gagctgatta
      541 aggagaacat gcacatgaag ctgtacatgg agggcaccgt gaacaaccac cacttcaagt
      601 gcacatccga gggcgaaggc aagccctacg agggcaccca gaccatgaga atcaaggtgg
      661 tcgagggcgg ccctctcccc ttcgccttcg acatcctggc taccagcttc atgtacggca
      721 gcagaacctt catcaaccac acccagggca tccccgactt ctttaagcag tccttccctg
      781 agggcttcac atgggagaga gtcaccacat acgaggacgg gggcgtgctg accgctaccc
      841 aggacaccag cctccaggac ggctgcctca tctacaacgt caagatcaga ggggtgaact
      901 tcccatccaa cggccctgtg atgcagaaga aaacactcgg ctgggaggcc aacaccgaga
      961 tgctgtaccc cgctgacggc ggcctggaag gcagaagcga catggccctg aagctcgtgg
     1021 gcgggggcca cctgatctgc aacttcaaga ccacatacag atccaagaaa cccgctaaga
     1081 acctcaagat gcccggcgtc tactatgtgg accacagact ggaaagaatc aaggaggccg
     1141 acaaagagac ctacgtcgag cagcacgagg tggctgtggc cagatactgc gacctcccta
     1201 gcaaactggg gcacaaactt aattgataaa ttccagtctt cggaaccaaa tcggatcccg
     1261 ggcccgtcga ctgcagaggc ctgcatgcaa gcttggtgta atcatggtca tagctgtttc
     1321 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     1381 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     1441 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     1501 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     1561 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     1621 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     1681 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     1741 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     1801 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     1861 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     1921 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     1981 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     2041 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     2101 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     2161 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     2221 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     2281 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     2341 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     2401 tccttttaaa ttaaaaatga agttttaaat caagcccaat ctgaataatg ttacaaccaa
     2461 ttaaccaatt ctgattagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata
     2521 tcaggattat caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca
     2581 ccgaggcagt tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca
     2641 acatcaatac aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca
     2701 ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc tttccagact
     2761 tgttcaacag gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta
     2821 ttcattcgtg attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta
     2881 caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca
     2941 cctgaatcag gatattcttc taatacctgg aatgctgttt ttccggggat cgcagtggtg
     3001 agtaaccatg catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat
     3061 tccgtcagcc agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg
     3121 ccatgtttca gaaacaactc tggcgcatcg ggcttcccat acaagcgata gattgtcgca
     3181 cctgattgcc cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg
     3241 gaatttaatc gcggcctcga cgtttcccgt tgaatatggc tcataacacc ccttgtatta
     3301 ctgtttatgt aagcagacag ttttattgtt catgatgata tatttttatc ttgtgcaatg
     3361 taacatcaga gattttgaga cacgggccag agctgca
//
