LOCUS       sequence-211164-        8059 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral expression of cytoplasmic TurboRFP with Blasticidin 
            selection in cells including neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8059)
  AUTHORS   Parker SS, Moutal A, Cai S, Chandrasekaran S, Roman MR, Koshy AA, 
            Khanna R, Zinsmaier KE, Mouneimne G
  TITLE     High Fidelity Cryopreservation and Recovery of Primary Rodent 
            Cortical Neurons.
  JOURNAL   eNeuro. 2018 Sep 27;5(5). pii: eN-MNT-0135-18. doi: 
            10.1523/ENEURO.0135-18.2018. eCollection 2018 Sep-Oct.
  PUBMED    30263951
REFERENCE   2  (bases 1 to 8059)
  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: "eNeuro."; 
            date: "2018-09-27"; volume: "5(5). pii: eN-MNT-0135-18. doi"; pages:
            " 10.1523/ENEURO.0135-18.2018. eCollection 2018 Sep-Oct"
COMMENT     Sequence Label: pLenti TurboRFP BlastR
FEATURES             Location/Qualifiers
     source          1..8059
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        15..241
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             242..422
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    469..594
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1087..1320
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1505..1549
                     /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    1816..1932
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2022..2225
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2176..2196
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2222..2246
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             2309..3004
                     /codon_start=1
                     /product="red fluorescent protein from Entacmaea 
                     quadricolor"
                     /label=TurboRFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMKIKV
                     VEGGPLPFAFDILATSFMYGSKAFINHTQGIPDFFKQSFPEGFTWERITTYEDGGVLTA
                     TQDTSFQNGCIIYNVKINGVNFPSNGPVMQKKTRGWEANTEMLYPADGGLRGHSQMALK
                     LVGGGYLHCSFKTTYRSKKPAKNLKMPGFHFVDHRLERIKEADKETYVEQHEMAVAKYC
                     DLPSKLGHR"
     ncRNA           3017..3110
                     /label=5' miR-30a
                     /note="sequence upstream of the 71-nt precursor of the 
                     human miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     misc_feature    join(3133..3707,3708..3710,3711..3719)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3300..3317)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3527..3546
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3726..4124
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    4147..4735
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4200..4220)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4618..4629)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4809..5042
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5114..5235
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5151..5170)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5205..5224
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5275..5410
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5443..5459)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5443..5459)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5456..5478)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5467..5483
                     /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(5491..5497,5498..5515,5516..5521))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5536..5557
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5674..5691)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5845..6433)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(6604..7395,7396..7464))
                     /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     7227..7246
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7465..7569)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7637..7655
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7693..7715)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7815..7834
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8028..8050
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     8042..8059
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     8043..8059
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gccaagctga cgcgtgtagt cttatgcaat actcttgtag tcttgcaaca tggtaacgat
       61 gagttagcaa catgccttac aaggagagaa aaagcaccgt gcatgccgat tggtggaagt
      121 aaggtggtac gatcgtgcct tattaggaag gcaacagacg ggtctgacat ggattggacg
      181 aaccactgaa ttgccgcatt gcagagatat tgtatttaag tgcctagctc gatacataaa
      241 cgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc
      301 actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt
      361 gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag
      421 cagtggcgcc cgaacaggga cttgaaagcg aaagggaaac cagaggagct ctctcgacgc
      481 aggactcggc ttgctgaagc gcgcacggca agaggcgagg ggcggcgact ggtgagtacg
      541 ccaaaaattt tgactagcgg aggctagaag gagagagatg ggtgcgagag cgtcagtatt
      601 aagcggggga gaattagatc gcgatgggaa aaaattcggt taaggccagg gggaaagaaa
      661 aaatataaat taaaacatat agtatgggca agcagggagc tagaacgatt cgcagttaat
      721 cctggcctgt tagaaacatc agaaggctgt agacaaatac tgggacagct acaaccatcc
      781 cttcagacag gatcagaaga acttagatca ttatataata cagtagcaac cctctattgt
      841 gtgcatcaaa ggatagagat aaaagacacc aaggaagctt tagacaagat agaggaagag
      901 caaaacaaaa gtaagaccac cgcacagcaa gcggccactg atcttcagac ctggaggagg
      961 agatatgagg gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc
     1021 attaggagta gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt
     1081 gggaatagga gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc
     1141 gtcaatgacg ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa
     1201 caatttgctg agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat
     1261 caagcagctc caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct
     1321 ggggatttgg ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag
     1381 ttggagtaat aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag
     1441 agaaattaac aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca
     1501 agaaaagaat gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt
     1561 taacataaca aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt
     1621 aggtttaaga atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc
     1681 accattatcg tttcagaccc acctcccaac cccgagggga cccgacaggc ccgaaggaat
     1741 agaagaagaa ggtggagaga gagacagaga cagatccatt cgattagtga acggatctcg
     1801 acggtatcgg ttaactttta aaagaaaagg ggggattggg gggtacagtg caggggaaag
     1861 aatagtagac ataatagcaa cagacataca aactaaagaa ttacaaaaac aaattacaaa
     1921 attcaaaatt ttatcgatac tagtattatg cccagtacat gaccttatgg gactttccta
     1981 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
     2041 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
     2101 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
     2161 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtt tatataagca
     2221 gagctcgttt agtgaaccgt cagatcgcct ggagacgcca tccacgctgt tttgacctcc
     2281 atagaagatt ctaggaccgg tcgccaccat gagcgagctg atcaaggaga acatgcacat
     2341 gaagctgtac atggagggca ccgtgaacaa ccaccacttc aagtgcacat ccgagggcga
     2401 aggcaagccc tacgagggca cccagaccat gaagatcaag gtggtcgagg gcggccctct
     2461 ccccttcgcc ttcgacatcc tggctaccag cttcatgtac ggcagcaaag ccttcatcaa
     2521 ccacacccag ggcatccccg acttctttaa gcagtccttc cctgagggct tcacatggga
     2581 gagaatcacc acatacgaag acgggggcgt gctgaccgct acccaggaca ccagcttcca
     2641 gaacggctgc atcatctaca acgtcaagat caacggggtg aacttcccat ccaacggccc
     2701 tgtgatgcag aagaaaacac gcggctggga ggccaacacc gagatgctgt accccgctga
     2761 cggcggcctg agaggccaca gccagatggc cctgaagctc gtgggcgggg gctacctgca
     2821 ctgctccttc aagaccacat acagatccaa gaaacccgct aagaacctca agatgcccgg
     2881 cttccacttc gtggaccaca gactggaaag aatcaaggag gccgacaaag agacctacgt
     2941 cgagcagcac gagatggctg tggccaagta ctgcgacctc cctagcaaac tggggcacag
     3001 ataatcgata gtttgtttga atgaggcttc agtactttac agaatcgttg cctgcacatc
     3061 ttggaaacac ttgctgggat tacttcttca ggttaaccca acagaaggct cgagcatgca
     3121 tctagagatc cgcccctctc cctccccccc ccctaacgtt actggccgaa gccgcttgga
     3181 ataaggccgg tgtgcgtttg tctatatgtt attttccacc atattgccgt cttttggcaa
     3241 tgtgagggcc cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc
     3301 tctcgccaaa ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc
     3361 ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg
     3421 cgacaggtgc ctctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca
     3481 accccagtgc cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tctcctcaag
     3541 cgtattcaac aaggggctga aggatgccca gaaggtaccc cattgtatgg gatctgatct
     3601 ggggcctcgg tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaaac gtctaggccc
     3661 cccgaaccac ggggacgtgg ttttcctttg aaaaacacga tgataagctt gccacaacca
     3721 ccgggatggc caagcctttg tctcaagaag aatccaccct cattgaaaga gcaacggcta
     3781 caatcaacag catccccatc tctgaagact acagcgtcgc cagcgcagct ctctctagcg
     3841 acggccgcat cttcactggt gtcaatgtat atcattttac tgggggacct tgtgcagaac
     3901 tcgtggtgct gggcactgct gctgctgcgg cagctggcaa cctgacttgt atcgtcgcga
     3961 tcggaaatga gaacaggggc atcttgagcc cctgcggacg gtgccgacag gtgcttctcg
     4021 atctgcatcc tgggatcaaa gccatagtga aggacagtga tggacagccg acggcagttg
     4081 ggattcgtga attgctgccc tctggttatg tgtgggaggg ctaagcgcgg ccgcggcgga
     4141 gtcgacaatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat
     4201 gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct
     4261 tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag
     4321 gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc
     4381 cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc
     4441 ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct
     4501 cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg
     4561 ctgctcgcct gtgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg
     4621 gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg
     4681 cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcctggt
     4741 acctttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt taaaagaaaa
     4801 ggggggactg gaagggctaa ttcactccca acgaagataa gatctgcttt ttgcttgtac
     4861 tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc
     4921 actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt
     4981 gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag
     5041 cagtagtagt tcatgtcatc ttattattca gtatttataa cttgcaaaga aatgaatatc
     5101 agagagtgag aggaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     5161 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     5221 catcaatgta tcttatcatg tctggctcta gctatcccgc ccctaactcc gcccatcccg
     5281 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     5341 tatgcagagg ccgaggcggc ctcggcctct gagctattcc agaagtagtg aggaggcttt
     5401 tttggaggcc tagacttttg cagagaccaa attcgtaatc atgtcatagc tgtttcctgt
     5461 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     5521 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     5581 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     5641 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     5701 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     5761 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     5821 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     5881 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     5941 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     6001 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     6061 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     6121 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     6181 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     6241 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     6301 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     6361 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     6421 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     6481 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     6541 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     6601 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     6661 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     6721 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     6781 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     6841 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     6901 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     6961 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     7021 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     7081 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     7141 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     7201 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     7261 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     7321 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     7381 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     7441 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     7501 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     7561 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat
     7621 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtctcgcgc gtttcggtga
     7681 tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc
     7741 ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg
     7801 ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata tgcggtgtga
     7861 aataccgcac agatgcgtaa ggagaaaata ccgcatcagg cgccattcgc cattcaggct
     7921 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
     7981 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
     8041 ttgtaaaacg acggccagt
//
