LOCUS       sequence-270305-        6601 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral expression of GFP-Ska1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6601)
  AUTHORS   Monda JK, Whitney IP, Tarasovetc EV, Wilson-Kubalek E, Milligan RA, 
            Grishchuk EL, Cheeseman IM
  TITLE     Microtubule Tip Tracking by the Spindle and Kinetochore Protein Ska1
            Requires Diverse Tubulin-Interacting Surfaces.
  JOURNAL   Curr Biol. 2017 Dec 4;27(23):3666-3675.e6. doi: 
            10.1016/j.cub.2017.10.018. Epub 2017 Nov 16.
  PUBMED    29153323
REFERENCE   2  (bases 1 to 6601)
  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.cub.2017.10.018"; journalName: "Curr Biol"; date: 
            "2017-12-4- 4"; volume: "27"; issue: "23"; pages: "3666-3675.e6"
COMMENT     Sequence Label: pIW093
FEATURES             Location/Qualifiers
     source          1..6601
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    100..299
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             500..916
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     846..868
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     884..900
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      951..960
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(957..959,960..962,963..1673)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(957..959,960..962,963..1673)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1002..1023)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1263..1282)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1610..1631
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1719..1739
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1752..1796
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2744..2764)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2749..3078
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2929..3064
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3086..3484
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     LTR             3653..4122
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(4252..4269)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4423..5011)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4503..4522)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5171..5962,5963..6031))
                     /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     5794..5813
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6032..6136)
                     /gene="bla"
                     /label=AmpR promoter
     LTR             join(6167..6601,1..35)
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
ORIGIN
        1 tgagtgattg actacccgtc agcgggggtc tttcatttgg gggctcgtcc gggatcggga
       61 gacccctgcc cagggaccac cgacccacca ccgggaggca agctggccag caacttatct
      121 gtgtctgtcc gattgtctag tgtctatgac tgattttatg cgcctgcgtc ggtactagtt
      181 agctaactag ctctgtatct ggcggacccg tggtggaact gacgagttct gaacacccgg
      241 ccgcaaccct gggagacgtc ccagggactt tgggggccgt ttttgtggcc cgacctgagg
      301 aagggagtcg atgtggaatc cgaccccgtc aggatatgtg gttctggtag gagacgagaa
      361 cctaaaacag ttcccgcctc cgtctgaatt tttgctttcg gtttggaacc gaagccgcgc
      421 gtcttgtctg ctgcagcgct gcagcatcgt tctgtgttgt ctctgtctga ctgtgtttct
      481 gtatttgtct gaaaattagg gccagactgt taccactccc ttaagtttga ccttaggtca
      541 ctggaaagat gtcgagcgga tcgctcacaa ccagtcggta gatgtcaaga agagacgttg
      601 ggttaccttc tgctctgcag aatggccaac ctttaacgtc ggatggccgc gagacggcac
      661 ctttaaccga gacctcatca cccaggttaa gatcaaggtc ttttcacctg gcccgcatgg
      721 acacccagac caggtcccct acatcgtgac ctgggaagcc ttggcttttg acccccctcc
      781 ctgggtcaag ccctttgtac accctaagcc tccgcctcct cttcctccat ccgccccgtc
      841 tctccccctt gaacctcctc gttcgacccc gcctcgatcc tccctttatc cagccctcac
      901 tccttctcta ggcgccggcc ggatcccagt gtggtggtac gctaccggtc gccaccatgg
      961 tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg
     1021 acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca
     1081 agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg
     1141 tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcagc
     1201 acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca
     1261 aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga
     1321 accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc
     1381 tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca
     1441 tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc
     1501 actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc
     1561 tgagcaccca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc
     1621 tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagtccggag
     1681 gggatcggtg gagctccacc ggtggcggcc gctctagaga gaatctttat tttcagggcg
     1741 ccgccaaatt caaagaaacc gctgctgcta aattcgaacg ccagcacatg gacagcggag
     1801 gtggaggtac tagtctcgag atggcaagca gcgacctgga gcagctgtgc tctcatgtta
     1861 atgaaaagat tggcaatatt aagaaaacct tatcattaag aaactgtgga caagagccca
     1921 cactgaagac cgtattaaat aaaataggag atgagatcat tgtaataaat gaacttctaa
     1981 ataaattgga attggaaatt cagtatcaag aacaaaccaa caattcactc aaggaactct
     2041 gtgaatctct tgaagaagat tacaaagaca tagaacatct taaagaaaac gttccttccc
     2101 atttgcctca agtaacagta acccagagct gtgttaaggg atcagatctt gatcctgaag
     2161 aaccaatcaa agttgaagaa cctgaacccg taaagaagcc tcccaaagag caaagaagta
     2221 ttaaggaaat gccatttata acttgtgatg agtttaacgg ggtgccaagt tacatgaaat
     2281 cggccctgac atacaaccag attaatgatg ttattaaaga aatcaacaag gcagtaatta
     2341 gtaaatataa aatcctacat cagccaaaaa agtctatgaa ctctgtgacc agaaatctct
     2401 atcacagatt tattgatgaa gaaacgaagg ataccaaagg tcgttatttc atcgtagagg
     2461 cagatattaa ggagttcaca actttgaaag ctgacaagaa gtttcacgtg ttactgaata
     2521 ttttacgaca ctgccggagg ctatcagagg tccgaggggg aggtttaaca cgatacgtaa
     2581 taacctgacc gcgggcccgg gatccaccgg atctagataa ctgatcataa tcagccatac
     2641 cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa
     2701 acataaaatg aatgcaattc gccagcacag tggtcgaccc tgtggaatgt gtgtcagtta
     2761 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     2821 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2881 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     2941 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     3001 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     3061 ggcctaggct tttgcaaaaa gcttcatggc caagcctttg tctcaagaag aatccaccct
     3121 cattgaaaga gcaacggcta caatcaacag catccccatc tctgaagact acagcgtcgc
     3181 cagcgcagct ctctctagcg acggccgcat cttcactggt gtcaatgtat atcattttac
     3241 tgggggacct tgtgcagaac tcgtggtgct gggcactgct gctgctgcgg cagctggcaa
     3301 cctgacttgt atcgtcgcga tcggaaatga gaacaggggc atcttgagcc cctgcggacg
     3361 gtgccgacag gtgcttctcg atctgcatcc tgggatcaaa gccatagtga aggacagtga
     3421 tggacagccg acggcagttg ggattcgtga attgctgccc tctggttatg tgtgggaggg
     3481 ctaagatcga taaaataaaa gattttattt agtctccaga aaaagggggg aatgaaagac
     3541 cccacctgta ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc atggaaaaat
     3601 acataactga gaatagagaa gttcagatca aggtcaggaa cagatggaac agctgaatat
     3661 gggccaaaca ggatatctgt ggtaagcagt tcctgccccg gctcagggcc aagaacagat
     3721 ggaacagctg aatatgggcc aaacaggata tctgtggtaa gcagttcctg ccccggctca
     3781 gggccaagaa cagatggtcc ccagatgcgg tccagccctc agcagtttct agagaaccat
     3841 cagatgtttc cagggtgccc caaggacctg aaatgaccct gtgccttatt tgaactaacc
     3901 aatcagttcg cttctcgctt ctgttcgcgc gcttctgctc cccgagctca ataaaagagc
     3961 ccacaacccc tcactcgggg cgccagtcct ccgattgact gagtcgcccg ggtacccgtg
     4021 tatccaataa accctcttgc agttgcatcc gacttgtggt ctcgctgttc cttgggaggg
     4081 tctcctctga gtgattgact acccgtcagc gggggtcttt cacatgcagc atgtatcaaa
     4141 attaatttgg ttttttttct taagtattta cattaaatgg ccatagttgc attaatgaat
     4201 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     4261 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     4321 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     4381 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     4441 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     4501 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     4561 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     4621 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     4681 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     4741 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     4801 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     4861 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     4921 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     4981 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     5041 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     5101 gatcttcacc tagatccttt tgcggccgca aatcaatcta aagtatatat gagtaaactt
     5161 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5221 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5281 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     5341 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5401 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5461 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     5521 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     5581 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     5641 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     5701 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     5761 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     5821 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     5881 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     5941 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     6001 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     6061 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6121 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgcagcctga atatgggcca
     6181 aacaggatat ctgtggtaag cagttcctgc cccggctcag ggccaagaac agatggaaca
     6241 gctgaatatg ggccaaacag gatatctgtg gtaagcagtt cctgccccgg ctcagggcca
     6301 agaacagatg gtccccagat gcggtccagc cctcagcagt ttctagagaa ccatcagatg
     6361 tttccagggt gccccaagga cctgaaatga ccctgtgcct tatttgaact aaccaatcag
     6421 ttcgcttctc gcttctgttc gcgcgcttct gctccccgag ctcaataaaa gagcccacaa
     6481 cccctcactc ggggcgccag tcctccgatt gactgagtcg cccgggtacc cgtgtatcca
     6541 ataaaccctc ttgcagttgc atccgacttg tggtctcgct gttccttggg agggtctcct
     6601 c
//
