LOCUS       sequence-240212-        8335 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral overexpression of Scara3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8335)
  AUTHORS   Henriksson J, Chen X, Gomes T, Ullah U, Meyer KB, Miragaia R, Duddy 
            G, Pramanik J, Yusa K, Lahesmaa R, Teichmann SA
  TITLE     Genome-wide CRISPR Screens in T Helper Cells Reveal Pervasive 
            Crosstalk between Activation and Differentiation.
  JOURNAL   Cell. 2019 Jan 7. pii: S0092-8674(18)31569-1. doi: 
            10.1016/j.cell.2018.11.044.
  PUBMED    30639098
REFERENCE   2  (bases 1 to 8335)
  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: "Cell. 2019 
            Jan 7. pii: S0092-8674(18)31569-1. doi: 10.1016/j.cell.2018.11.044."
COMMENT     Sequence Label: MSCV-Scara3-IRES-BFP
FEATURES             Location/Qualifiers
     source          1..8335
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..517
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     misc_feature    581..922
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             989..1405
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1335..1357
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1373..1389
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     RBS             2673..2681
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     misc_feature    3298..3847
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3442..3459)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3669..3688
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(3850..3852,3853..3855,3856..4569)
                     /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(3895..3916)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     LTR             4730..5244
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5406..5422
                     /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(5430..5436,5437..5454,5455..5460))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5475..5496
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5613..5630)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5784..6372)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5864..5883)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6543..7334,7335..7403))
                     /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     7166..7185
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7404..7508)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7576..7594
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7632..7654)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7754..7773
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7967..7989
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     8117..8136
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 aatgaaagac cccacctgta ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc
       61 atggaaaata cataactgag aatagagaag ttcagatcaa ggttaggaac agagagacag
      121 cagaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc tcagggccaa
      181 gaacagatgg tccccagatg cggtcccgcc ctcagcagtt tctagagaac catcagatgt
      241 ttccagggtg ccccaaggac ctgaaatgac cctgtgcctt atttgaacta accaatcagt
      301 tcgcttctcg cttctgttcg cgcgcttctg ctccccgagc tcaataaaag agcccacaac
      361 ccctcactcg gcgcgccagt cctccgatag actgcgtcgc ccgggtaccc gtattcccaa
      421 taaagcctct tgctgtttgc atccgaatcg tggactcgct gatccttggg agggtctcct
      481 cagattgatt gactgcccac ctcgggggtc tttcatttgg aggttccacc gagatttgga
      541 gacccctgcc cagggaccac cgaccccccc gccgggaggt aagctggcca gcggtcgttt
      601 cgtgtctgtc tctgtctttg tgcgtgtttg tgccggcatc taatgtttgc gcctgcgtct
      661 gtactagtta gctaactagc tctgtatctg gcggacccgt ggtggaactg acgagttctg
      721 aacacccggc cgcaaccctg ggagacgtcc cagggacttt gggggccgtt tttgtggccc
      781 gacctgagga agggagtcga tgtggaatcc gaccccgtca ggatatgtgg ttctggtagg
      841 agacgagaac ctaaaacagt tcccgcctcc gtctgaattt ttgctttcgg tttggaaccg
      901 aagccgcgcg tcttgtctgc tgcagcgctg cagcatcgtt ctgtgttgtc tctgtctgac
      961 tgtgtttctg tatttgtctg aaaattaggg ccagactgtt accactccct taagtttgac
     1021 cttaggtcac tggaaagatg tcgagcggat cgctcacaac cagtcggtag atgtcaagaa
     1081 gagacgttgg gttaccttct gctctgcaga atggccaacc tttaacgtcg gatggccgcg
     1141 agacggcacc tttaaccgag acctcatcac ccaggttaag atcaaggtct tttcacctgg
     1201 cccgcatgga cacccagacc aggtccccta catcgtgacc tgggaagcct tggcttttga
     1261 cccccctccc tgggtcaagc cctttgtaca ccctaagcct ccgcctcctc ttcctccatc
     1321 cgccccgtct ctcccccttg aacctcctcg ttcgaccccg cctcgatcct ccctttatcc
     1381 agccctcact ccttctctag gcgccggaat tcagatctta cgtagctagc ggatcttaac
     1441 catgaaagtg agatcagctg gcagtgacag ggatgtactt tgcgtcacag aagaggactt
     1501 ggcaggagaa gatgaggata tgccttcttt tccttgcacc caagaaggta gggcagggcc
     1561 acgctgtaat aggtgccaaa aaaatctttc actccatacc tctgtccgga tattgtacct
     1621 gttcttgact cttctgttgg ttgccgttgc tgtgcttgct agccttgtgt tccggaaagt
     1681 cgacagcctc agtgaggata tctctctcgc ccagagtatt tataataaga agctggtcag
     1741 tatgcaagag aacttgcaag gccttgatcc caaagctctg ataaattgtt ccttctgtcg
     1801 ggaagccgaa caactcggac aggaaattcg aaaagtccag gaagagcttg agggcctgca
     1861 gaaaatgctg cttgcccaag aagtacaact tgatcaaaca tcacaaactc atgaacttct
     1921 ctccacccgg agttctcaaa tttcccagga gatgggcagc tgctcattta gcatacacca
     1981 agtaaaccag tctctgggcc ttttccttgc tcaagttaga gggtggcagg ctacaacagc
     2041 tggaatggat attactttga aagacctgac acaagagtgc tacgacgtaa aagccgctgt
     2101 ccatcagatc aatttcacag tgggacaaac agccgaatgg atacacggaa tacaacgcaa
     2161 aaccgatgag gaaacactta cattgcagaa gattgttact gactggcaga actatacacg
     2221 gctcttcggc ggactcagga ctaccagcgc aaagaccgga gaaatcgtaa agactattca
     2281 gactacactg ggggcctcta gtcaacgaat ctcccagaac agcgagtcaa tgcatgacct
     2341 ggtactgcaa gtgatgggac tgcagctgca gctggacaac atcagttcat tcctcgacga
     2401 tcatgaagag aatatgcatg atctccaata ccacacacgg tacgctcaaa atcgaacagt
     2461 tgagcgcttt gaaagcctgg agggccggat ggcaagccac gaaatagaga tagggacaat
     2521 tttcactaac ataaatgcta ctgataatca tgtgcactca atgctgaaat atttggatga
     2581 tgttcgcctt tcttgcaccc tcggttttca cactcacgcc gaggagttgt actatctgaa
     2641 taagtccgtc agtctgatgc tgggaacaac tgacctcctt agagaacgct tctcattgct
     2701 ctccgcaagg ctcgatttca atgtacggaa tctcagcatg attatggagg agatgaaggc
     2761 agtggataca catcacgggg agatcctgcg gaatgtcaca gttatccggg gcgtaccagg
     2821 tccacctggt ccaaggggcc ttaaatgtga tacaggtgtt aaaggacccg tgggatccag
     2881 aggtccaaaa ggtgacccag ggaatctggg gccacccgga ccccagggtc ctcaaggaca
     2941 gccaggggag cctggcccag taggggagcg aggccccgct gggcctcgag ggtttcccgg
     3001 actcaaagga tccaaaggct ctttcgggac aggaggacct aggggccagc ccggaccaaa
     3061 gggcgatgtt ggccccctcg gtcccgaggg cccccctggc agccccggtc cttcagggcc
     3121 tcaggggaaa ccagggataa gtggaaaaac aggaagccct ggacagcggg gagccactgg
     3181 acctaaaggt gaaccaggca tccaagggcc tcctgggctt ccaggccctc caggtccacc
     3241 agggaaccag agcccctact gataggctcg agcgggatca attccgcccc ccccctaacg
     3301 ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg ttattttcca
     3361 ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc ttcttgacga
     3421 gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg aatgtcgtga
     3481 aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg accctttgca
     3541 ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca cgtgtataag
     3601 atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata gttgtggaaa
     3661 gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc cagaaggtac
     3721 cccattgtat gggatctgat ctggggcctc ggtgcacatg ctttacatgt gtttagtcga
     3781 ggttaaaaaa acgtctaggc cccccgaacc acggggacgt ggttttcctt tgaaaaacac
     3841 gataatacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc
     3901 gagctggacg gcgacgtaaa cggccacaag ttcagcgtga ggggcgaggg cgagggcgat
     3961 gccaccaacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc
     4021 tggcccaccc tcgtgaccac cctgagccac ggcgtgcagt gcttcgcccg ctaccccgac
     4081 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc
     4141 accatcttct tcaaggacga cggcacctac aagacccgcg ccgaggtgaa gttcgagggc
     4201 gacaccctgg tgaaccgcat cgagctgaag ggcgtcgact tcaaggagga cggcaacatc
     4261 ctggggcaca agctggagta caacttcaac agccacaaca tctatatcat ggccgtcaag
     4321 cagaagaacg gcatcaaggt gaacttcaag atccgccaca acgtggagga cggcagcgtg
     4381 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc
     4441 gacagccact acctgagcac ccagtccgtg ctgagcaaag accccaacga gaagcgcgat
     4501 cacatggtcc tgctggagtt ccgcaccgcc gccgggatca ctctcggcat ggacgagctg
     4561 tacaagtaag cggccgcgac tctagagtcg acctgcaggc atgcaagctt caggtagccg
     4621 gctaacgtta acaaccggta cctctagaac tatagctagc atgcgcaaat ttaaagcgct
     4681 gatatcgata aaataaaaga ttttatttag tctccagaaa aaggggggaa tgaaagaccc
     4741 cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat ggaaaataca
     4801 taactgagaa tagagaagtt cagatcaagg ttaggaacag agagacagca gaatatgggc
     4861 caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga acagatggtc
     4921 cccagatgcg gtcccgccct cagcagtttc tagagaacca tcagatgttt ccagggtgcc
     4981 ccaaggacct gaaatgaccc tgtgccttat ttgaactaac caatcagttc gcttctcgct
     5041 tctgttcgcg cgcttctgct ccccgagctc aataaaagag cccacaaccc ctcactcggc
     5101 gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt gtatccaata aaccctcttg
     5161 cagttgcatc cgacttgtgg tctcgctgtt ccttgggagg gtctcctctg agtgattgac
     5221 tacccgtcag cgggggtctt tcatgggtaa cagtttcttg aagttggaga acaacattct
     5281 gagggtagga gtcgaatatt aagtaatcct gactcaatta gccactgttt tgaatccaca
     5341 tactccaata ctcctgaaat agttcattat ggacagcgca gaaagagctg gggagaattg
     5401 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     5461 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     5521 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     5581 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     5641 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     5701 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     5761 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     5821 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     5881 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     5941 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     6001 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     6061 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     6121 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     6181 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     6241 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     6301 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     6361 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     6421 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     6481 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     6541 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     6601 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     6661 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     6721 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     6781 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     6841 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     6901 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     6961 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     7021 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     7081 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     7141 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     7201 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     7261 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     7321 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     7381 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     7441 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     7501 gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg
     7561 acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat
     7621 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     7681 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     7741 tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat gcggtgtgaa
     7801 ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc attcaggctg
     7861 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa
     7921 gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt
     7981 tgtaaaacga cggcgcaagg aatggtgcat gcaaggagat ggcgcccaac agtcccccgg
     8041 ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag
     8101 cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc
     8161 cggtgatgcc ggccacgatg cgtccggcgt agaggcgatt agtccaattt gttaaagaca
     8221 ggatatcagt ggtccaggct ctagttttga ctcaacaata tcaccagctg aagcctatag
     8281 agtacgagcc atagataaaa taaaagattt tatttagtct ccagaaaaag ggggg
//
