LOCUS       sequence-149456-        7163 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor vector for 3' FLAG tag of human HHEX .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7163)
  TITLE     Mendenhall CRISPR tagging plasmids
REFERENCE   2  (bases 1 to 7163)
  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: pFETCh_HHEX
FEATURES             Location/Qualifiers
     source          1..7163
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             847..870
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             874..897
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             901..924
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             940..996
                     /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             1009..1803
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1063..1082)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1673..1692
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(2665..2681)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2674..2696)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(2900..3715)
                     /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="MSHIQRETSCSKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3623..3642
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3707..3726
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4095..4683
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4584..4603
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4837..4854
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5154..5175
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5190..5195,5196..5213,5214..5220)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5228..5244
                     /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     5233..5255
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5252..5268
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5252..5268
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        5445..5647
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5598..5618
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5644..5668
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             5744..6874
                     /codon_start=1
                     /product="herpes simplex virus thymidine kinase"
                     /label=HSV TK
                     /note="confers sensitivity to nucleoside analogs such as 
                     acyclovir or ganciclovir"
                     /translation="MASYPCHQHASAFDQAARSRGHSNRRTALRPRRQQEATEVRLEQK
                     MPTLLRVYIDGPHGMGKTTTTQLLVALGSRDDIVYVPEPMTYWQVLGASETIANIYTTQ
                     HRLDQGEISAGDAAVVMTSAQITMGMPYAVTDAVLAPHIGGEAGSSHAPPPALTLIFDR
                     HPIAALLCYPAARYLMGSMTPQAVLAFVALIPPTLPGTNIVLGALPEDRHIDRLAKRQR
                     PGERLDLAMLAAIRRVYGLLANTVRYLQGGGSWREDWGQLSGTAVPPQGAEPQSNAGPR
                     PHIGDTLFTLFRAPELLAPNGDLYNVFAWALDVLAKRLRPMHVFILDYDQSPAGCRDAL
                     LQLTSGMVQTHVTTPGSIPTICDLARTFAREMGEAN"
ORIGIN
        1 cagttggcag cattcttcca gatccacatt ttttttcttt gccatgattt tgtaacttgt
       61 taaacattta cacatgtaga tgtttataaa tttactccag aaaatgaatg aaaaaagatc
      121 cagtctaata attgaaactt tattcttaat tcaccagtta gcattcttat ttttaaaatg
      181 tctgaaagac tgttaattgg caaggtttaa atatttaaag aatgaagtta aggtgtatct
      241 gtagtgatct gattttcttt cccccttaaa cttttgagaa tattgaaacc atgggtctgg
      301 gctggtgtta agtaaccaga atttgtgttc aattggtata atgtatctgc ccttatgttc
      361 atgactggtt ctcatctttt aagcatataa tctttgatat gacagaaact tttcttccaa
      421 agtgagcaaa aagcacaagt atgtatctcc tgaataagat tataaactga aaacactcag
      481 taaactaaaa attaaaagat ggaataggat gaagagacat tttgatttat tcctctcacc
      541 tatccatatt ttatgatcct tccaatctcc attttcattt cctttattca ggagaaccct
      601 caaagcaata aaaaagaaga actggaaagt ttggacagtt cctgtgatca gaggcaagat
      661 ttgcccagtg aacagaataa aggtgcttct ttggatagct ctcaatgttc gccctcccct
      721 gcctcccagg aagaccttga atcagagatt tcagaggatt ctgatcagga agtggacatt
      781 gagggcgata aaagctattt taatgctgga gggagcggag gaggttccgg tggaggtggt
      841 tctggagatt acaaggatga cgacgataag ggcgattaca aggatgacga cgataaggga
      901 gattacaagg atgacgacga taaggtttca ggaagcggag ctactaactt cagcctgctg
      961 aagcaggctg gagacgtgga ggagaaccct ggacctggat cgtttcgcat gattgaacaa
     1021 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     1081 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     1141 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     1201 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     1261 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     1321 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     1381 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     1441 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     1501 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     1561 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     1621 ggattcatcg actgtggccg gctgggtgtg gccgaccgct atcaggacat agcgttggct
     1681 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     1741 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     1801 tgattcgaac atccattggc atgttcagaa aactggattt aggaataatg ttttgctaca
     1861 gaaaatcttc atagaagaac tggaaggcta tataagaaag ggaatcaatt ctctggtatt
     1921 ctggaaacct aaaaatattt ggtgcactgc tcaattaaca aacctacatg gagaccttaa
     1981 ttttgactta acaaatagtt tatgtactgc tcttaggttg ttttgataaa gtgacattat
     2041 agtgattaaa ttcttccccc tttaaaaaaa cagttagtgg ttttcactat ttataaaaaa
     2101 ttaattttga actttttgtt aaatttttaa gttatagctt taaaggtttt aataggacct
     2161 tcttgaacga cttttctgta atctgtttat ctcccactta atggaaaggc aaaggggtac
     2221 cccaaatcca gaggtgccta catttcaggc agccttggag tattttaaaa ggaaaacatt
     2281 ctttactttt atatgacatt cttatactgc tgtctcaaat ccaaaaacat ttcagagctc
     2341 ttgtctcaga gatgtgtgtt ctttttgtca gagatatggt tgatgagaat cttaaatgct
     2401 tgttttgcac tatcacttag tacctgtttg accaaggtgt taaggggata gtacctccca
     2461 attcaagcag agaaactgac ctgactaaag ttaatcgcag atgaactaga agtcacaggt
     2521 tattaaatgt aagtagattg tagatactgt tttatatcaa acaatgttta taatgtgtat
     2581 atagaattgt tcactgtaaa aaaaatggcc aacttggaaa gtcctctcca ctgactgtag
     2641 cctccaattc actggccgtc gttcactggc cgtcgtttta caacgtcgtg actgggaaaa
     2701 ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa
     2761 tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg
     2821 cgatttattc aacaaagccg ccgtcccgtc aagtcagcgt aatgctctgc cagtgttaca
     2881 accaattaac caattctgat tagaaaaact catcgagcat caaatgaaac tgcaatttat
     2941 tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa
     3001 actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc
     3061 gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga
     3121 aatcaccatg agtgacgact gaatccggtg agaatggcaa aagcttatgc atttctttcc
     3181 agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac
     3241 cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac
     3301 aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat
     3361 tttcacctga atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag
     3421 tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca
     3481 taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac
     3541 ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg
     3601 tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca
     3661 tgttggaatt taatcgcggc ttcgagcaag acgtttcccg ttgaatatgg ctcataacac
     3721 cccttgtatt actgtttatg taagcagaca gttttattgt tcatgatgat atatttttat
     3781 cttgtgcaat gtaacatcag agattttgag acacaacgtg gctttgttga ataaatcgaa
     3841 cttttgctga gttgaaggat cagatcacgc atcttcccga caacgcagac cgttccgtgg
     3901 caaagcaaaa gttcaaaatc accaactggt ccacctacaa caaagctctc atcaaccgtg
     3961 gctccctcac tttctggctg gatgatgggg cgattcaggc ctggtatgag tcagcaacac
     4021 cttcttcacg aggcagacct ctcgacggag ttccactgag cgtcagaccc cgtagaaaag
     4081 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     4141 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     4201 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     4261 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     4321 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     4381 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     4441 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     4501 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     4561 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     4621 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     4681 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     4741 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     4801 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     4861 gaagaagctc gcacattcag cagcgttttt cagcgcgttt tcgatcaacg tttcaatgtt
     4921 ggtatcaaca ccaggtttaa ctttgaactt atcggcactg acggttactg attttgaact
     4981 tttgctttgc cacggaacgg tctgcgttgt cgggaagatg cgtgatctga tccttcaact
     5041 cagcaaaagt tcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc
     5101 agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg
     5161 agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg
     5221 tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc
     5281 aagcttgcat gcctgcaggt cgactctaga ggatccccgg gtaccgagct cgaattagct
     5341 tgcatgcctg caggtcaatt ccctggcatt atgcccagta catgacctta tgggactttc
     5401 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
     5461 agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttcaagtc tccaccccat
     5521 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
     5581 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
     5641 cagagctcgt ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct
     5701 ccataggaga caccgggacc gatccagcct ggggatcccg accatggctt cgtacccctg
     5761 ccatcaacac gcgtctgcgt tcgaccaggc tgcgcgttct cgcggccata gcaaccgacg
     5821 tacggcgttg cgccctcgcc ggcagcaaga agccacggaa gtccgcctgg agcagaaaat
     5881 gcccacgcta ctgcgggttt atatagacgg tcctcacggg atggggaaaa ccaccaccac
     5941 gcaactgctg gtggccctgg gttcgcgcga cgatatcgtc tacgtacccg agccgatgac
     6001 ttactggcag gtgctggggg cttccgagac aatcgcgaac atctacacca cacaacaccg
     6061 cctcgaccag ggtgagatat cggccgggga cgcggcggtg gtaatgacaa gcgcccagat
     6121 aacaatgggc atgccttatg ccgtgaccga cgccgttctg gctcctcata tcggggggga
     6181 ggctgggagc tcacatgccc cgcccccggc cctcaccctc atcttcgacc gccatcccat
     6241 cgccgccctc ctgtgctacc cggccgcgcg ataccttatg ggcagcatga ccccccaggc
     6301 cgtgctggcg ttcgtggccc tcatcccgcc gaccttgccc ggcacaaaca tcgtgttggg
     6361 ggcccttccg gaggacagac acatcgaccg cctggccaaa cgccagcgcc ccggcgagcg
     6421 gcttgacctg gctatgctgg ccgcgattcg ccgcgtttac gggctgcttg ccaatacggt
     6481 gcggtatctg cagggcggcg ggtcgtggcg ggaggattgg ggacagcttt cggggacggc
     6541 cgtgccgccc cagggtgccg agccccagag caacgcgggc ccacgacccc atatcgggga
     6601 cacgttattt accctgtttc gggcccccga gttgctggcc cccaacggcg acctgtacaa
     6661 cgtgtttgcc tgggccttgg acgtcttggc caaacgcctc cgtcccatgc acgtctttat
     6721 cctggattac gaccaatcgc ccgccggctg ccgggacgcc ctgctgcaac ttacctccgg
     6781 gatggtccag acccacgtca ccacccccgg ctccataccg acgatctgcg acctggcgcg
     6841 cacgtttgcc cgggagatgg gggaggctaa ctgaggatcc cacgtcacta ttgtatactc
     6901 tatattatac tctatgttat actctgtaat cctactcaat aaacgtgtca cgcctgtgaa
     6961 accgtactaa gtctcccgtg tattcttatc accatcaggt gacatcctcg cccaggctgt
     7021 caatcatgcc ggtatcgatt ccagtagcac cggccccacg ctgacaaccc actcttgcag
     7081 cgttagcagc gcccctctta acaagccgac ccccaccagc gtcgcggtta ctaacactcc
     7141 tctccccgac ctgcagccca gct
//
