LOCUS       sequence-44618-e        7615 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7615)
  TITLE     Tsoulfas Lab plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7615)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pLV-EBFP2-nuc
FEATURES             Location/Qualifiers
     source          1..7615
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    97..685
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(150..170)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(568..579)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             772..1005
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    1077..1198
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1114..1133)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1168..1187
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      1238..1373
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1393..1412)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1394..1412)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1422..1439)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1422..1438)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1431..1453)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1580..2035
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1667..1686)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1877..1898
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2061..2165
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2166..2234,2235..3026)
                     /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(2384..2403)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3197..3785
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3686..3705
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3939..3956
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4073..4094
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4109..4114,4115..4132,4133..4139)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4147..4163
                     /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     4152..4174
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4171..4187
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4171..4187
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     4206..4226
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        4208..4226
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        4254..4480
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             4481..4661
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    4708..4833
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5326..5559
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             5744..5788
                     /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    6055..6171
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        6194..6497
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6498..6701
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6652..6672
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     6698..6722
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      6777..6786
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(6783..6785,6786..6788,6789..7499)
                     /codon_start=1
                     /product="enhanced blue variant of GFP (Ai et al., 2007)"
                     /label=EBFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLSHGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGVDFKEDGNILGHKLEYNFNSHNIYIMAVKQKNGIK
                     VNFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDSHYLSTQSVLSKDPNEKRDHMVLL
                     EFRTAAGITLGMDELYK"
     primer_bind     complement(6828..6849)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             7524..7544
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7548..7568
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7572..7592
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
ORIGIN
        1 gatcccaccg gtgctcgaga tgcatactag tgaattcacg cgttctagac ctgcagggtc
       61 gactccggat gatcagggcc ctgtacagat atcgacaatc aacctctgga ttacaaaatt
      121 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct
      181 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg
      241 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc
      301 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt
      361 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc
      421 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg
      481 ttgtcgggga agctgacgtc ctttccatgg ctgctcgcct gtgttgccac ctggattctg
      541 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc
      601 ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg
      661 atctcccttt gggccgcctc cccgcctgga attcgagctc ggtaccttta agaccaatga
      721 cttacaaggc agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc
      781 taattcactc ccaacgaaga caagatctgc tttttgcttg tactgggtct ctctggttag
      841 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat
      901 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact
      961 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtagt agttcatgtc
     1021 atcttattat tcagtattta taacttgcaa agaaatgaat atcagagagt gagaggaact
     1081 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     1141 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     1201 atgtctggct ctagctatcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     1261 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     1321 cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctagggac
     1381 gtacccaatt cgccctatag tgagtcgtat tacgcgcgct cactggccgt cgttttacaa
     1441 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
     1501 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     1561 agcctgaatg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     1621 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     1681 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     1741 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     1801 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     1861 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     1921 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     1981 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg
     2041 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     2101 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     2161 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     2221 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     2281 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     2341 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     2401 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     2461 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     2521 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     2581 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     2641 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     2701 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     2761 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     2821 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     2881 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     2941 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     3001 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     3061 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     3121 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     3181 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     3241 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     3301 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     3361 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     3421 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     3481 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     3541 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     3601 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     3661 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     3721 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     3781 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     3841 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
     3901 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
     3961 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     4021 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
     4081 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
     4141 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
     4201 agcgcgcaat taaccctcac taaagggaac aaaagctgga gctgcaagct taatgtagtc
     4261 ttatgcaata ctcttgtagt cttgcaacat ggtaacgatg agttagcaac atgccttaca
     4321 aggagagaaa aagcaccgtg catgccgatt ggtggaagta aggtggtacg atcgtgcctt
     4381 attaggaagg caacagacgg gtctgacatg gattggacga accactgaat tgccgcattg
     4441 cagagatatt gtatttaagt gcctagctcg atacataaac gggtctctct ggttagacca
     4501 gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag
     4561 cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag
     4621 atccctcaga cccttttagt cagtgtggaa aatctctagc agtggcgccc gaacagggac
     4681 ttgaaagcga aagggaaacc agaggagctc tctcgacgca ggactcggct tgctgaagcg
     4741 cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt gactagcgga
     4801 ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag aattagatcg
     4861 cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt aaaacatata
     4921 gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt agaaacatca
     4981 gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg atcagaagaa
     5041 cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag gatagagata
     5101 aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag taagaccacc
     5161 gcacagcaag cggccgctga tcttcagacc tggaggagga gatatgaggg acaattggag
     5221 aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag cacccaccaa
     5281 ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag ctttgttcct
     5341 tgggttcttg ggagcagcag gaagcactat gggcgcagcg tcaatgacgc tgacggtaca
     5401 ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga gggctattga
     5461 ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc aggcaagaat
     5521 cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg gttgctctgg
     5581 aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata aatctctgga
     5641 acagatttgg aatcacacga cctggatgga gtgggacaga gaaattaaca attacacaag
     5701 cttaatacac tccttaattg aagaatcgca aaaccagcaa gaaaagaatg aacaagaatt
     5761 attggaatta gataaatggg caagtttgtg gaattggttt aacataacaa attggctgtg
     5821 gtatataaaa ttattcataa tgatagtagg aggcttggta ggtttaagaa tagtttttgc
     5881 tgtactttct atagtgaata gagttaggca gggatattca ccattatcgt ttcagaccca
     5941 cctcccaacc ccgaggggac ccgacaggcc cgaaggaata gaagaagaag gtggagagag
     6001 agacagagac agatccattc gattagtgaa cggatctcga cggtatcggt taacttttaa
     6061 aagaaaaggg gggattgggg ggtacagtgc aggggaaaga atagtagaca taatagcaac
     6121 agacatacaa actaaagaat tacaaaaaca aattacaaaa ttcaaaattt tatcgataag
     6181 cttgggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
     6241 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
     6301 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
     6361 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
     6421 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
     6481 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
     6541 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
     6601 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
     6661 gcggtaggcg tgtacggtgg gaggtctata taagcagagc tcgtttagtg aaccgtcaga
     6721 tcgcctggag acgccatcca cgctgttttg acctccatag aagacaccgc ccggtcgcca
     6781 ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
     6841 acggcgacgt aaacggccac aagttcagcg tgaggggcga gggcgagggc gatgccacca
     6901 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
     6961 ccctcgtgac caccctgagc cacggcgtgc agtgcttcgc ccgctacccc gaccacatga
     7021 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
     7081 tcttcaagga cgacggcacc tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
     7141 tagtgaaccg catcgagctg aagggcgtcg acttcaagga ggacggcaac atcctggggc
     7201 acaagctgga gtacaacttc aacagccaca acatctatat catggccgtc aagcagaaga
     7261 acggcatcaa ggtgaacttc aagatccgcc acaacgtgga ggacggcagc gtgcagctcg
     7321 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacagcc
     7381 actacctgag cacccagtcc gtgctgagca aagaccccaa cgagaagcgc gatcacatgg
     7441 tcctgctgga gttccgcacc gccgccggga tcactctcgg catggacgag ctgtacaagt
     7501 ccggactcag atctcgagct gatccaaaaa agaagagaaa ggtagatcca aaaaagaaga
     7561 gaaaggtaga tccaaaaaag aagagaaagg taggatccac cggatctaga taact
//
