LOCUS       sequence-257265-        7369 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses CMV promoter driving puromycin resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7369)
  AUTHORS   Hartenian EN, Glaunsinger BA
  TITLE     Xrn1 activity broadly represses RNA polymerase II occupancy at 
            mammalian but not viral promoters during herpesvirus infection
  JOURNAL   bioRxiv 585984
REFERENCE   2  (bases 1 to 7369)
  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: "bioRxiv 
            585984"
COMMENT     Sequence Label: pLKO-CMV-puro
FEATURES             Location/Qualifiers
     source          1..7369
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             176..220
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    441..558
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        700..1079
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1080..1283
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1234..1254
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             1290..1889
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1290..1309)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1786..1806
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    1907..2495
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1960..1980)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2378..2389)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             2567..2800
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2878..2999
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2915..2934)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2969..2988
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      3039..3174
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3194..3213)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3195..3213)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3223..3240)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3223..3239)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3232..3254)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3381..3836
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3468..3487)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3678..3699
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3862..3966
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3967..4035,4036..4827)
                     /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     complement(4185..4204)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4998..5586
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5487..5506
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5740..5757
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5874..5895
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5910..5915,5916..5933,5934..5940)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5948..5964
                     /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     5953..5975
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5972..5988
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5972..5988
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6007..6027
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6009..6027
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6055..6281
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6282..6462
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6509..6634
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7127..7360
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
ORIGIN
        1 gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta gttggagtaa
       61 taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca gagaaattaa
      121 caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc aagaaaagaa
      181 tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt ttaacataac
      241 aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg taggtttaag
      301 aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt caccattatc
      361 gtttcagacc cacctcccaa ccccgactcg aaagcttggc gtaactagat cttgagacaa
      421 atggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg
      481 gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt
      541 acaaaaattc aaaattttcg ggtttattac agggacagca gagatccact ttggcgccgg
      601 ctcgaggggg cccgggtgca aagatggata aagttttaaa cagagaggaa tctttgcagc
      661 taatggacct tctaggtctt gaaaggagtg ggaatcgatg acattgatta ttgactagtt
      721 attaatagta atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta
      781 cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt
      841 caataatgac gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg
      901 tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagta
      961 cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga
     1021 ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg
     1081 tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc
     1141 caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact
     1201 ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt
     1261 gggaggtcta tataagcaga gctggtacca tgaccgagta caagcccacg gtgcgcctcg
     1321 ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc gccgactacc
     1381 ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc gagctgcaag
     1441 aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg
     1501 ccgccgtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga
     1561 tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa cagatggaag
     1621 gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc gtcggagtct
     1681 cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg
     1741 ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac ctccccttct
     1801 acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct
     1861 ggtgcatgac ccgcaagccc ggtgcctgat gtacaagtaa gtcgacaatc aacctctgga
     1921 ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg
     1981 tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt
     2041 ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag
     2101 gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc
     2161 caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga
     2221 actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa
     2281 ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct gtgttgccac
     2341 ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct
     2401 tccttcccgc ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca
     2461 gacgagtcgg atctcccttt gggccgcctc cccgcgtcga ctttaagacc aatgacttac
     2521 aaggcagctg tagatcttag ccacttttta aaagaaaagg ggggactgga agggctaatt
     2581 cactcccaac gaagacaaga tctgcttttt gcttgtactg ggtctctctg gttagaccag
     2641 atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc
     2701 ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga
     2761 tccctcagac ccttttagtc agtgtggaaa atctctagca gtacgtatag tagttcatgt
     2821 catcttatta ttcagtattt ataacttgca aagaaatgaa tatcagagag tgagaggaac
     2881 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     2941 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     3001 catgtctggc tctagctatc ccgcccctaa ctccgcccat cccgccccta actccgccca
     3061 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     3121 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggga
     3181 cgtacccaat tcgccctata gtgagtcgta ttacgcgcgc tcactggccg tcgttttaca
     3241 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc
     3301 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg
     3361 cagcctgaat ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     3421 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     3481 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     3541 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     3601 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     3661 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     3721 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     3781 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt
     3841 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     3901 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     3961 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     4021 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     4081 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     4141 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     4201 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     4261 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     4321 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     4381 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     4441 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     4501 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     4561 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     4621 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     4681 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     4741 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     4801 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     4861 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     4921 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     4981 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     5041 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     5101 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     5161 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     5221 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     5281 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     5341 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     5401 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     5461 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     5521 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     5581 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     5641 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     5701 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     5761 gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc
     5821 agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg
     5881 agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg
     5941 tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc
     6001 aagcgcgcaa ttaaccctca ctaaagggaa caaaagctgg agctgcaagc ttaatgtagt
     6061 cttatgcaat actcttgtag tcttgcaaca tggtaacgat gagttagcaa catgccttac
     6121 aaggagagaa aaagcaccgt gcatgccgat tggtggaagt aaggtggtac gatcgtgcct
     6181 tattaggaag gcaacagacg ggtctgacat ggattggacg aaccactgaa ttgccgcatt
     6241 gcagagatat tgtatttaag tgcctagctc gatacataaa cgggtctctc tggttagacc
     6301 agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa
     6361 gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga
     6421 gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc cgaacaggga
     6481 cttgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc ttgctgaagc
     6541 gcgcacggca agaggcgagg ggcggcgact ggtgagtacg ccaaaaattt tgactagcgg
     6601 aggctagaag gagagagatg ggtgcgagag cgtcagtatt aagcggggga gaattagatc
     6661 gcgatgggaa aaaattcggt taaggccagg gggaaagaaa aaatataaat taaaacatat
     6721 agtatgggca agcagggagc tagaacgatt cgcagttaat cctggcctgt tagaaacatc
     6781 agaaggctgt agacaaatac tgggacagct acaaccatcc cttcagacag gatcagaaga
     6841 acttagatca ttatataata cagtagcaac cctctattgt gtgcatcaaa ggatagagat
     6901 aaaagacacc aaggaagctt tagacaagat agaggaagag caaaacaaaa gtaagaccac
     6961 cgcacagcaa gcggccgctg atcttcagac ctggaggagg agatatgagg gacaattgga
     7021 gaagtgaatt atataaatat aaagtagtaa aaattgaacc attaggagta gcacccacca
     7081 aggcaaagag aagagtggtg cagagagaaa aaagagcagt gggaatagga gctttgttcc
     7141 ttgggttctt gggagcagca ggaagcacta tgggcgcagc gtcaatgacg ctgacggtac
     7201 aggccagaca attattgtct ggtatagtgc agcagcagaa caatttgctg agggctattg
     7261 aggcgcaaca gcatctgttg caactcacag tctggggcat caagcagctc caggcaagaa
     7321 tcctggctgt ggaaagatac ctaaaggatc aacagctcct ggggatttg
//
