LOCUS       sequence-431543-        3073 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fragmid fragment: (2A Selection) Blasticidin resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3073)
  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 3073)
  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: AA006
FEATURES             Location/Qualifiers
     source          1..3073
                     /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..497
                     /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             504..902
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     primer_bind     complement(983..999)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(983..999)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(996..1018)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1007..1023
                     /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(1031..1037,1038..1055,1056..1061))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1076..1097
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1214..1231)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1385..1973)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1465..1484)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2151..2960)
                     /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     2874..2893
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2952..2971
                     /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 agcgagggca gaggaagcct tctaacatgc ggtgacgtgg
      481 aggagaatcc cggcccttcc gggatggcca agcctttgtc tcaagaagaa tccaccctca
      541 ttgaaagagc aacggctaca atcaacagca tccccatctc tgaggactac agcgtcgcca
      601 gcgcagctct ctctagcgac ggccgcatct tcactggtgt caatgtatat cattttactg
      661 ggggaccttg tgcagaactc gtggtgctgg gcactgctgc tgctgcggca gctggcaacc
      721 tgacttgtat cgtcgcgatc ggaaatgaga acaggggcat cttgagcccc tgcggacggt
      781 gccgacaggt gcttctcgat ctgcatcctg ggatcaaagc catagtgaag gacagtgatg
      841 gacagccgac ggcagttggg attcgtgaat tgctgccctc tggttatgtg tgggagggct
      901 aataaattcc agtcttcgga accaaatcgg atcccgggcc cgtcgactgc agaggcctgc
      961 atgcaagctt ggtgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     1021 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     1081 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     1141 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     1201 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     1261 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     1321 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     1381 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     1441 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     1501 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     1561 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     1621 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     1681 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     1741 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     1801 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     1861 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     1921 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     1981 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     2041 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     2101 ttaaatcaag cccaatctga ataatgttac aaccaattaa ccaattctga ttagaaaaac
     2161 tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag gattatcaat accatatttt
     2221 tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga ggcagttcca taggatggca
     2281 agatcctggt atcggtctgc gattccgact cgtccaacat caatacaacc tattaatttc
     2341 ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat gagtgacgac tgaatccggt
     2401 gagaatggca aaagtttatg catttctttc cagacttgtt caacaggcca gccattacgc
     2461 tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg
     2521 agacgaaata cgcgatcgct gttaaaagga caattacaaa caggaatcga atgcaaccgg
     2581 cgcaggaaca ctgccagcgc atcaacaata ttttcacctg aatcaggata ttcttctaat
     2641 acctggaatg ctgtttttcc ggggatcgca gtggtgagta accatgcatc atcaggagta
     2701 cggataaaat gcttgatggt cggaagaggc ataaattccg tcagccagtt tagtctgacc
     2761 atctcatctg taacatcatt ggcaacgcta cctttgccat gtttcagaaa caactctggc
     2821 gcatcgggct tcccatacaa gcgatagatt gtcgcacctg attgcccgac attatcgcga
     2881 gcccatttat acccatataa atcagcatcc atgttggaat ttaatcgcgg cctcgacgtt
     2941 tcccgttgaa tatggctcat aacacccctt gtattactgt ttatgtaagc agacagtttt
     3001 attgttcatg atgatatatt tttatcttgt gcaatgtaac atcagagatt ttgagacacg
     3061 ggccagagct gca
//
