LOCUS       sequence-375676-        6361 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral delivery of control guide RNA .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6361)
  AUTHORS   Hosokawa H, Ungerback J, Wang X, Matsumoto M, Nakayama KI, Cohen SM,
            Tanaka T, Rothenberg EV
  TITLE     Transcription Factor PU.1 Represses and Activates Gene Expression in
            Early T Cells by Redirecting Partner Transcription Factor Binding.
  JOURNAL   Immunity. 2018 Jun 19;48(6):1119-1134.e7. doi: 
            10.1016/j.immuni.2018.04.024.
  PUBMED    29924977
REFERENCE   2  (bases 1 to 6361)
  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"; doi: 
            "10.1016/j.immuni.2018.04"; journalName: "Immunity"; date: 
            "2018-06-19- 19"; volume: "48"; issue: "6"; pages: "1119-1134.e7"
COMMENT     Sequence Label: E42_gControl_dTet_IRES_mTurquoise2
FEATURES             Location/Qualifiers
     source          1..6361
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      6..141
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     177..196
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(277..294)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(448..1036)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(528..547)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1207..1998,1999..2067))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     1830..1849
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2068..2172)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2240..2258
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        2358..2737
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2738..2941
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2892..2912
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             2942..3117
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    3180..3379
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             3580..3996
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     3926..3948
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     3964..3980
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        4012..4260
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     4012..4032
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     4183..4202
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        4400..4611
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      4628..4637
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4634..4636,4637..4639,4640..5353)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Goedhart et 
                     al., 2012)"
                     /label=mTurquoise2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYFSDNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4679..4700)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4940..4959)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5287..5308
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             5496..5921
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(6024..6046)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6182..6202)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        join(6187..6361,1..155)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
ORIGIN
        1 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
       61 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
      121 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct tactggctta actatgcggc
      181 atcagagcag attgtactga gagtgcacca tatgcggtgt gaaataccgc acagatgcgt
      241 aaggagaaaa taccgcatca ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
      301 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
      361 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
      421 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
      481 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
      541 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
      601 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
      661 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
      721 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
      781 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
      841 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg
      901 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
      961 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     1021 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     1081 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     1141 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     1201 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     1261 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     1321 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     1381 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     1441 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     1501 gcgcaacgtt gttgccattg ctgcaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     1561 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     1621 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     1681 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     1741 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     1801 gagttgctct tgcccggcgt caacacggga taataccgcg ccacatagca gaactttaaa
     1861 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     1921 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     1981 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     2041 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     2101 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     2161 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat
     2221 catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtcttc aagaattagc
     2281 ttggccattg catacgttgt atccatatca taatatgtac atttatattg gctcatgtcc
     2341 aacattaccg ccatgttgac attgattatt gactagttat taatagtaat caattacggg
     2401 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
     2461 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
     2521 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
     2581 ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga
     2641 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg
     2701 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat
     2761 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
     2821 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc
     2881 cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
     2941 tcaataaaag agcccacaac ccctcactcg gcgcgccagt cttccgatag actgcgtcgc
     3001 ccgggtaccc gtattcccaa taaagcctct tgctgtttgc atccgaatcg tggtctcgct
     3061 gttccttggg agggtctcct ctgagtgatt gactacccac gacgggggtc tttcatttgg
     3121 gggctcgtcc gggatttgga gacccctgcc cagggaccac cgacccacca ccgggaggta
     3181 agctggccag caacttatct gtgtctgtcc gattgtctag tgtctatgtt tgatgttatg
     3241 cgcctgcgtc tgtactagtt agctaactag ctctgtatct ggcggacccg tggtggaact
     3301 gacgagttct gaacacccgg ccgcaaccct gggagacgtc ccagggactt tgggggccgt
     3361 ttttgtggcc cgacctgagg aagggagtcg atgtggaatc cgaccccgtc aggatatgtg
     3421 gttctggtag gagacgagaa cctaaaacag ttcccgcctc cgtctgaatt tttgctttcg
     3481 gtttggaacc gaagccgcgc gtcttgtctg ctgcagcgct gcagcatcgt tctgtgttgt
     3541 ctctgtctga ctgtgtttct gtatttgtct gaaaattagg gccagactgt taccactccc
     3601 ttaagtttga ccttaggtca ctggaaagat gtcgagcgga tcgctcacaa ccagtcggta
     3661 gatgtcaaga agagacgttg ggttaccttc tgctctgcag aatggccaac ctttaacgtc
     3721 ggatggccgc gagacggcac ctttaaccga gacctcatca cccaggttaa gatcaaggtc
     3781 ttttcacctg gcccgcatgg acacccagac caggtcccct acatcgtgac ctgggaagcc
     3841 ttggcttttg acccccctcc ctgggtcaag ccctttgtac accctaagcc tccgcctcct
     3901 cttcctccat ccgccccgtc tctccccctt gaacctcctc gttcgacccc gcctcgatcc
     3961 tccctttatc cagccctcac tccttctcta ggcgccggaa ttgaaggatc cgagggccta
     4021 tttcccatga ttccttcata tttgcatata cgatacaagg ctgttagaga gataattgga
     4081 attaatttga ctgtaaacac aaagatatta gtacaaaata cgtgacgtag aaagtaataa
     4141 tttcttgggt agtttgcagt tttaaaatta tgttttaaaa tggactatca tatgcttacc
     4201 gtaacttgaa agtatttcga tttcttggct ttatatatct tgtggaaagg acgaaacacc
     4261 gggcatttcg cagcctaccg gtttaagagc tatgctggaa acagcatagc aagtttaaat
     4321 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt tttctcgagt
     4381 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
     4441 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
     4501 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
     4561 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtgtcgtga
     4621 cgcggtcgcc accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
     4681 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
     4741 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     4801 gccctggccc accctcgtga ccaccctgtc ctggggcgtg cagtgcttcg cccgctaccc
     4861 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     4921 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
     4981 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     5041 catcctgggg cacaagctgg agtacaacta ctttagcgac aacgtctata tcaccgccga
     5101 caagcagaag aacggcatca aggccaactt caagatccgc cacaacatcg aggacggcgg
     5161 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     5221 gcccgacaac cactacctga gcacccagtc caagctgagc aaagacccca acgagaagcg
     5281 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     5341 gctgtacaag taaatcgatt agtccaattt gttaaagaca ggatatcagt ggtccaggct
     5401 ctagttttga ctcaacaata tcaccagctg aagcctatag agtacgagcc atagataaaa
     5461 taaaagattt tatttagtct ccagaaaaag gggggaatga aagaccccac ctgtaggttt
     5521 ggcaagctag cttaagtaac gccattttgc aaggcatgga aaaatacata actgagaata
     5581 gagaagttca gatcaaggtc aggaacagat ggaacagggt cgaccctaga gaaccatcag
     5641 atgtttccag ggtgccccaa ggacctgaaa tgaccctgtg ccttatttga actaaccaat
     5701 cagttcgctt ctcgcttctg ttcgcgcgct tctgctcccc gagctcaata aaagagccca
     5761 caacccctca ctcggggcgc cagtcctccg attgactgag tcgcccgggt acccgtgtat
     5821 ccaataaacc ctcttgcagt tgcatccgac ttgtggtctc gctgttcctt gggagggtct
     5881 cctctgagtg attgactacc cgtcagcggg ggtctttcat ttgggggctc gtccgggatc
     5941 gggagacccc tgcccaggga ccaccgaccc accaccggga ggtaagctgg ctgcctcgcg
     6001 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     6061 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     6121 gggtgtcggg gcgcagccat gacccagtca cgtagcgata gcggagtgta gatccggctg
     6181 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     6241 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     6301 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     6361 c
//
