LOCUS       sequence-132115-        7153 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor vector for 3' FLAG tag of human DNMT3B.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7153)
  TITLE     Mendenhall CRISPR tagging plasmids
REFERENCE   2  (bases 1 to 7153)
  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_DNMT3B
FEATURES             Location/Qualifiers
     source          1..7153
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             837..860
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             864..887
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             891..914
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             930..986
                     /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             999..1793
                     /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(1053..1072)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1663..1682
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(2655..2671)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2664..2686)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(2890..3705)
                     /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     3613..3632
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3697..3716
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4085..4673
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4574..4593
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4827..4844
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5144..5165
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5180..5185,5186..5203,5204..5210)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5218..5234
                     /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     5223..5245
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5242..5258
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5242..5258
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        5435..5637
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5588..5608
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5634..5658
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             5734..6864
                     /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 aagaccaagc aagacacagg cctgaggtat cacatctttg gcatgtttct gaataaccca
       61 cgcagccatt ggcctgttgg gatgactttt agtattggat tgtatacctg agtaacactg
      121 aggctcagcc tctgtagcct ctttccaggc ctggtagaat cctatagagg aagcttttgg
      181 tatatttgat gccatctgcc tggagtcatc tagcatggtc cttcatccgg cagctgggtt
      241 tgaagaggga aggaaaactc tggttgcctg agcctgtgat gactgaaaat ctaggctgtt
      301 tttgagtgac cttgttcctt attaaggaaa ttgcttaact tctcccatca tagtgttttg
      361 aaaatggata cattgctggc tggctgattt agtatgcatg catttattct aggaaggtcc
      421 tctgagcacc aactctatgc caagagtatg gtgctaggcc tgcacaaaac gggggacttt
      481 catttgtgag caatttgtaa gcaagtgttc tgaattaagg gctctgaatt taggtccttg
      541 gggaccttac tgatgggact gagggatggc gagggcagaa agagtgggac ctggctggtt
      601 gaggctgtca acatcctgga ggcacttctg acttgctgtc ttttcactcc ggtaccccca
      661 ggatctttgg ctttcctgtg cactacacag acgtgtccaa catgggccgt ggtgcccgcc
      721 agaagctgct gggaaggtcc tggagcgtgc ctgtcatccg acacctcttc gcccctctga
      781 aggactactt tgcatgtgaa gggagcggag gaggttccgg tggaggtggt tctggagatt
      841 acaaggatga cgacgataag ggcgattaca aggatgacga cgataaggga gattacaagg
      901 atgacgacga taaggtttca ggaagcggag ctactaactt cagcctgctg aagcaggctg
      961 gagacgtgga ggagaaccct ggacctggat cgtttcgcat gattgaacaa gatggattgc
     1021 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     1081 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     1141 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
     1201 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     1261 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     1321 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     1381 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     1441 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     1501 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     1561 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     1621 actgtggccg gctgggtgtg gccgaccgct atcaggacat agcgttggct acccgtgata
     1681 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     1741 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgattcgaac
     1801 atcgccaggc cccaagccca ctggggtgtg tggcagagcc aggacccagg aggtgtgatt
     1861 cctgaaggca tccccaggcc ctgctcttcc tcagctgtgt gggtcatacc gtgtacctca
     1921 gttccctctt gctcagtggg ggcagagcca cctgactctt gcaggggtag cctgaggtgc
     1981 cgcctccttg tgcacaaatc agacctggct gcttggagca gcctaacacg gtgctcattt
     2041 tttcttctcc taaaacttta aaacttgaag taggtagcaa cgtggctttt tttttttccc
     2101 ttcctgggtc taccactcag agaaacaatg gctaagatac caaaaccaca gtgccgacag
     2161 ctctccaata ctcaggttaa tgctgaaaaa tcatccaaga cagttattgc aagagtttaa
     2221 tttttgaaaa ctggctactg ctctgtgttt acagacgtgt gcagttgtag gcatgtagct
     2281 acaggacatt tttaagggcc caggatcgtt ttttcccagg gcaagcagaa gagaaaatgt
     2341 tgtatatgtc ttttacccgg cacattcccc ttgcctaaat acaagggctg gagtctgcac
     2401 gggacctatt agagtatttt ccacaatgat gatgatttca gcagggatga cgtcatcatc
     2461 acattcaggg ctattttttc ccccacaaac ccaagggcag gggccactct tagctaaatc
     2521 cctccccgtg actgcaatag aaccctctgg ggagctcagg aaggggtgtg ctgagttcta
     2581 taatataagc tgccatatat ttcttggaaa gtcctctcca ctgactgtag cctccaattc
     2641 actggccgtc gttcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
     2701 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
     2761 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg cgatttattc
     2821 aacaaagccg ccgtcccgtc aagtcagcgt aatgctctgc cagtgttaca accaattaac
     2881 caattctgat tagaaaaact catcgagcat caaatgaaac tgcaatttat tcatatcagg
     2941 attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa actcaccgag
     3001 gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc gtccaacatc
     3061 aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg
     3121 agtgacgact gaatccggtg agaatggcaa aagcttatgc atttctttcc agacttgttc
     3181 aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac cgttattcat
     3241 tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac aattacaaac
     3301 aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat tttcacctga
     3361 atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag tggtgagtaa
     3421 ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca taaattccgt
     3481 cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac ctttgccatg
     3541 tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg tcgcacctga
     3601 ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca tgttggaatt
     3661 taatcgcggc ttcgagcaag acgtttcccg ttgaatatgg ctcataacac cccttgtatt
     3721 actgtttatg taagcagaca gttttattgt tcatgatgat atatttttat cttgtgcaat
     3781 gtaacatcag agattttgag acacaacgtg gctttgttga ataaatcgaa cttttgctga
     3841 gttgaaggat cagatcacgc atcttcccga caacgcagac cgttccgtgg caaagcaaaa
     3901 gttcaaaatc accaactggt ccacctacaa caaagctctc atcaaccgtg gctccctcac
     3961 tttctggctg gatgatgggg cgattcaggc ctggtatgag tcagcaacac cttcttcacg
     4021 aggcagacct ctcgacggag ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     4081 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     4141 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     4201 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     4261 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     4321 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     4381 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     4441 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     4501 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     4561 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     4621 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     4681 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     4741 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     4801 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagaagctc
     4861 gcacattcag cagcgttttt cagcgcgttt tcgatcaacg tttcaatgtt ggtatcaaca
     4921 ccaggtttaa ctttgaactt atcggcactg acggttactg attttgaact tttgctttgc
     4981 cacggaacgg tctgcgttgt cgggaagatg cgtgatctga tccttcaact cagcaaaagt
     5041 tcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     5101 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca
     5161 ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg
     5221 tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc aagcttgcat
     5281 gcctgcaggt cgactctaga ggatccccgg gtaccgagct cgaattagct tgcatgcctg
     5341 caggtcaatt ccctggcatt atgcccagta catgacctta tgggactttc ctacttggca
     5401 gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa
     5461 tgggcgtgga tagcggtttg actcacgggg atttcaagtc tccaccccat tgacgtcaat
     5521 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
     5581 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt
     5641 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccataggaga
     5701 caccgggacc gatccagcct ggggatcccg accatggctt cgtacccctg ccatcaacac
     5761 gcgtctgcgt tcgaccaggc tgcgcgttct cgcggccata gcaaccgacg tacggcgttg
     5821 cgccctcgcc ggcagcaaga agccacggaa gtccgcctgg agcagaaaat gcccacgcta
     5881 ctgcgggttt atatagacgg tcctcacggg atggggaaaa ccaccaccac gcaactgctg
     5941 gtggccctgg gttcgcgcga cgatatcgtc tacgtacccg agccgatgac ttactggcag
     6001 gtgctggggg cttccgagac aatcgcgaac atctacacca cacaacaccg cctcgaccag
     6061 ggtgagatat cggccgggga cgcggcggtg gtaatgacaa gcgcccagat aacaatgggc
     6121 atgccttatg ccgtgaccga cgccgttctg gctcctcata tcggggggga ggctgggagc
     6181 tcacatgccc cgcccccggc cctcaccctc atcttcgacc gccatcccat cgccgccctc
     6241 ctgtgctacc cggccgcgcg ataccttatg ggcagcatga ccccccaggc cgtgctggcg
     6301 ttcgtggccc tcatcccgcc gaccttgccc ggcacaaaca tcgtgttggg ggcccttccg
     6361 gaggacagac acatcgaccg cctggccaaa cgccagcgcc ccggcgagcg gcttgacctg
     6421 gctatgctgg ccgcgattcg ccgcgtttac gggctgcttg ccaatacggt gcggtatctg
     6481 cagggcggcg ggtcgtggcg ggaggattgg ggacagcttt cggggacggc cgtgccgccc
     6541 cagggtgccg agccccagag caacgcgggc ccacgacccc atatcgggga cacgttattt
     6601 accctgtttc gggcccccga gttgctggcc cccaacggcg acctgtacaa cgtgtttgcc
     6661 tgggccttgg acgtcttggc caaacgcctc cgtcccatgc acgtctttat cctggattac
     6721 gaccaatcgc ccgccggctg ccgggacgcc ctgctgcaac ttacctccgg gatggtccag
     6781 acccacgtca ccacccccgg ctccataccg acgatctgcg acctggcgcg cacgtttgcc
     6841 cgggagatgg gggaggctaa ctgaggatcc cacgtcacta ttgtatactc tatattatac
     6901 tctatgttat actctgtaat cctactcaat aaacgtgtca cgcctgtgaa accgtactaa
     6961 gtctcccgtg tattcttatc accatcaggt gacatcctcg cccaggctgt caatcatgcc
     7021 ggtatcgatt ccagtagcac cggccccacg ctgacaaccc actcttgcag cgttagcagc
     7081 gcccctctta acaagccgac ccccaccagc gtcgcggtta ctaacactcc tctccccgac
     7141 ctgcagccca gct
//
