LOCUS       sequence-475906-        5989 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ScVip1-536-1107-K558A-K605A-K732A-K817A protein expression in insect
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5989)
  AUTHORS   Raia P, Lee K, Bartsch SM, Rico-Resendiz F, Portugal-Calisto D, 
            Vadas O, Panse VG, Fiedler D, Hothorn M
  TITLE     A small signaling domain controls PPIP5K phosphatase activity in 
            phosphate homeostasis.
  JOURNAL   Nat Commun. 2025 Feb 19;16(1):1753. doi: 10.1038/s41467-025-56937-0.
  PUBMED    39966396
REFERENCE   2  (bases 1 to 5989)
  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: "Nat 
            Commun."; date: "2025-02-19"; volume: "16(1):1753. doi"; pages: " 
            10.1038/s41467-025-56937-0"
COMMENT     Sequence Label: pBB3-ScVip1-536-1107-K558A-K605A-K732A-K817A
FEATURES             Location/Qualifiers
     source          1..5989
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     mobile_element  724..947
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(1014..1547)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(1736..1741,1742..1758,1759..1764))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     protein_bind    complement(1835..1868)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     polyA_signal    2123..2171
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     2196..2215
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(join(2260..2973,2974..2976,2977..2979))
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2305..2326)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2654..2673
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2913..2934
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     promoter        complement(3021..3130)
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     promoter        3175..3266
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     3191..3208
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             3311..3340
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             join(3350..3373,3374..3409,3410..3433)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="WSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     CDS             3449..3469
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     polyA_signal    5380..5514
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5417..5436)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5471..5490
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  5635..5800
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattgacttg ggtcgctctt cctgtggatg cgcagatgcc ctgcgtaagc
      721 gggtgtgggc ggacaataaa gtcttaaact gaacaaaata gatctaaact atgacaataa
      781 agtcttaaac tagacagaat agttgtaaac tgaaatcagt ccagttatgc tgtgaaaaag
      841 catactggac ttttgttatg gctaaagcaa actcttcatt ttctgaagtg caaattgccc
      901 gtcgtattaa agaggggcgt ggccaagggc atgtaaagac tatattcgcg gcgttgtgac
      961 aatttaccga acaactccgc ggccgggaag ccgatctcgg cttgaacgaa ttgttaggtg
     1021 gcggtacttg ggtcgatatc aaagtgcatc acttcttccc gtatgcccaa ctttgtatag
     1081 agagccactg cgggatcgtc accgtaatct gcttgcacgt agatcacata agcaccaagc
     1141 gcgttggcct catgcttgag gagattgatg agcgcggtgg caatgccctg cctccggtgc
     1201 tcgccggaga ctgcgagatc atagatatag atctcactac gcggctgctc aaacttgggc
     1261 agaacgtaag ccgcgagagc gccaacaacc gcttcttggt cgaaggcagc aagcgcgatg
     1321 aatgtcttac tacggagcaa gttcccgagg taatcggagt ccggctgatg ttgggagtag
     1381 gtggctacgt ctccgaactc acgaccgaaa agatcaagag cagcccgcat ggatttgact
     1441 tggtcagggc cgagcctaca tgtgcgaatg atgcccatac ttgagccacc taactttgtt
     1501 ttagggcgac tgccctgctg cgtaacatcg ttgctgctgc gtaacatcgt tgctgctcca
     1561 taacatcaaa catcgaccca cggcgtaacg cgcttgctgc ttggatgccc gaggcataga
     1621 ctgtacaaaa aaacagtcat aacaagccat gaaaaccgcc actgcgccgt taccaccgct
     1681 gcgttcggtc aaggttctgg accagttgcg tgagcgcata cgctacttgc attacagttt
     1741 acgaaccgaa caggcttatg tcaactgggt tcgtgccttc atccgtttcc acggtgtgcg
     1801 tcacccggca accttgggca gcagcgaagt cgccataact tcgtatagca tacattatac
     1861 gaagttatct gtaactataa cggtcctaag gtagcgagtt taaacactag tatcgattcg
     1921 cgacctactc cggaaggttt aaacgtaccc gtagtggcta tggcagggct tgccgccccg
     1981 acgttggctg cgagccctgg gccttcaccc gaacttgggg gttggggtgg ggaaaaggaa
     2041 gaaacgcggg cgtattggtc ccaatggggt ctcggtgggg tatcgacaga gtgccagccc
     2101 tgggaccgaa ccccgcgttt atgaacaaac gacccaacac ccgtgcgttt tattctgtct
     2161 ttttattgcc gtcatagcgc gggttccttc cggtattgtc tccttccgtg tttcagttag
     2221 cctcccccat ctcccggtac cgcatgctcc gattcgatcc tacttgtaca gctcgtccat
     2281 gccgagagtg atcccggcgg cggtcacgaa ctccagcagg accatgtgat cgcgcttctc
     2341 gttggggtct ttgctcaggg cggactggta gctcaggtag tggttgtcgg gcagcagcac
     2401 ggggccgtcg ccgatggggg tgttctgctg gtagtggtcg gcgagctgca cgctgccgtc
     2461 ctcgatgttg tggcggatct tgaagttcac cttgatgccg ttcttctgct tgtcggccat
     2521 gatatagacg ttgtggctgt tgtagttgta ctccagcttg tgccccagga tgttgccgtc
     2581 ctccttgaag tcgataccct tcagctcgat gcggttcacc agggtgtcgc cctcgaactt
     2641 cacctcggcg cgggtcttgt agttgccgtc gtccttgaag aagatggtgc gctcctggac
     2701 gtagccttcg ggcatggcgg acttgaagaa gtcgtgctgc ttcatgtggt cggggtagcg
     2761 ggcgaagcac tgcaggccgt agccgaaggt ggtcacgagg gtgggccagg gcacgggcag
     2821 cttgccggtg gtgcagatga acttcagggt cagcttgccg taggtggcat cgccctcgcc
     2881 ctcgccggac acgctgaact tgtggccgtt tacgtcgccg tccagctcga ccaggatggg
     2941 caccaccccg gtgaacagct cctcgccctt gctcaccatc atggtttcgg accgcccggg
     3001 tgatcaagtc ttcgtcgagt gattgtaaat aaaatgtaat ttacagtata gtattttaat
     3061 taatatacaa atgatttgat aataattctt atttaactat aatatattgt gttgggttga
     3121 attaaaggtc cgtatactag tatcgattcg cgacctactc cggaatatta atagatcatg
     3181 gagataatta aaatgataac catctcgcaa ataaataagt attttactgt tttcgtaaca
     3241 gttttgtaat aaaaaaacct ataaatattc cggattattc ataccgtccc accatcgggc
     3301 gcggatgaag catcatcatc atcatcacca tcaccaccac ggtgctagct ggtcccaccc
     3361 ccagttcgag aagggtggtg gttccggtgg cggatccggt ggttctgctt ggagccaccc
     3421 tcaattcgaa aagggctccg gttccggcga gaacctgtac ttccaaggtg ccaaatgggt
     3481 gttcaagggc ctcgccatca tcatccgtca cgctgaccgt actcctaagc aaaagttcgc
     3541 tcactccttc actagcccca tcttcatctc cctgctgaag ggtcacaaag aggaagtcgt
     3601 catccgtaac gtgaacgatc tgaagatcgt gctgcaggct ctgcgtatcg ctctggacga
     3661 gaaggctggc aaccccgctg ctatcaaggt gctggctaac gctctcgaga agaagctgaa
     3721 cttccccggc accaagatcc agctgaagcc agtgctgaac aaagagaacg aggtcgagaa
     3781 ggtgcagttc atcctgaagt ggggtggcga gcctactcac tccgctaagt accaggctac
     3841 cgagctggga gagcagatgc gtcaagactt cgacctgttg aacaagtcca tcctgcagaa
     3901 tatcaagatc ttcagctcct ccgagcgtcg tgtgctgcac actgctcagt actggacccg
     3961 tgctctgttc ggtgctgacg agctgggttc cgacgagatc tctatccgca aggacctgct
     4021 ggacgactcc aacgctgcta aggacctcat ggacaaagtc gctaagaagt tgaagcccct
     4081 gctgcgcgag ggcaaagagg ctcctcctca attcgcttgg ccctccaaga tgcccgagcc
     4141 ttacctggtc atcaagcgtg tggtcgagct gatgaactac cacaagaaaa tcatggataa
     4201 caacttcgcc aagaaagacg tgaactccat gcagacccgt tggtgcacct ctgaggaccc
     4261 cagcctgttc aaagagcgtt gggacaagct tttcaaagag ttcaacaacg ccgaggctgt
     4321 cgacccttcc aagatctccg agctgtacga caccatgaag tacgacgctc tgcacaaccg
     4381 tcagttcttg gagaacatct tcgaccccgg cctgcctaac gaggctatcg ctgacgaact
     4441 cggttctcac tccctggtgg atcgttaccc catcaacgtc ctggccaaga acaacttcaa
     4501 gatcatcgac tcccactcca tgaacaactc cggcaagaac tcctccaact ccgtgggttc
     4561 tctcggctgg gtgctcgagt ctggaaagac ctcaaccgct cgtaacccca agtcctctag
     4621 ccagttcgac gagccccgtt tcatgcagct gcgcgaactg tacaagctgg ctaaggtcct
     4681 gttcgacttc atctgcccca aagagtacgg tatctccgac gctgagaagc tggacatcgg
     4741 cctgctgact tctctgcctc tggctaagca gatcctgaac gacatcggcg acatgaagaa
     4801 ccgcgagact cccgcttgtg tggcttactt caccaaagag tcccacatct acaccctgct
     4861 caacatcatc tacgaaagcg gtatccccat gaggatcgct cgcaacgctc tgcccgaact
     4921 ggactacttg tcccagatca ccttcgagtt gtacgagtcc accgacgctt ccggccagaa
     4981 gtcccactct atccgtctga agatgtctcc cggttgccac actcaggacc ctctggacgt
     5041 gcaattggac gaccgtcact acatctcttg catccctaag atctccctga ccaagcacct
     5101 cgacatggac tacgtgcagc agaagctgcg taacaagttc acccgtgtga tcatgcctcc
     5161 aaagttcact cccgtgaaca tcacctcata agcggccgca ctcgagtagg atcgaatcgg
     5221 actttcgaat ctagagcctg cagtctcgac aagcttgtcg agaagtacta gaggatcata
     5281 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
     5341 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     5401 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     5461 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg gatctgatca
     5521 ctgcttgagc ctagaagatc cggctgctaa caaagcccga aaggaagctg agttggctgc
     5581 tgccaccgct gagcaataac tatcataacc cctagggtat acccatctaa ttggaaccag
     5641 ataagtgaaa tctagttcca aactattttg tcatttttaa ttttcgtatt agcttacgac
     5701 gctacaccca gttcccatct attttgtcac tcttccctaa ataatcctta aaaactccat
     5761 ttccacccct cccagttccc aactattttg tccgcccaca accggttgac ttgggtcaac
     5821 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     5881 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     5941 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttc
//
