LOCUS       sequence-478356-        5989 bp    DNA     circular SYN 14-JUL-2025
DEFINITION  ScVip1-536-1107-R547A-H548A-R551A 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 Jul 14, 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-R547A-H548A-R551A
FEATURES             Location/Qualifiers
     source          1..5989
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             7..36
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             join(46..69,70..105,106..129)
                     /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             145..165
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     polyA_signal    2076..2210
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2113..2132)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2167..2186
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  2331..2496
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      2686..3274
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3175..3194
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     mobile_element  3409..3632
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(3699..4232)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(4421..4426,4427..4443,4444..4449))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     protein_bind    complement(4520..4553)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     polyA_signal    4808..4856
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     4881..4900
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(join(4945..5658,5659..5661,5662..5664))
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4990..5011)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     5339..5358
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5598..5619
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     promoter        complement(5706..5815)
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     promoter        5860..5951
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     5876..5893
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
ORIGIN
        1 atgaagcatc atcatcatca tcaccatcac caccacggtg ctagctggtc ccacccccag
       61 ttcgagaagg gtggtggttc cggtggcgga tccggtggtt ctgcttggag ccaccctcaa
      121 ttcgaaaagg gctccggttc cggcgagaac ctgtacttcc aaggtgccaa atgggtgttc
      181 aagggcctcg ccatcatcat cgctgccgct gacgctactc ctaagcaaaa gttcaagcac
      241 tccttcacta gccccatctt catctccctg ctgaagggtc acaaagagga agtcgtcatc
      301 cgtaacgtga acgatctgaa gatcgtgctg caggctctgc gtatcgctct ggacgagaag
      361 gctggcaacc ccgctaagat caaggtgctg gctaacgctc tcgagaagaa gctgaacttc
      421 cccggcacca agatccagct gaagccagtg ctgaacaaag agaacgaggt cgagaaggtg
      481 cagttcatcc tgaagtgggg tggcgagcct actcactccg ctaagtacca ggctaccgag
      541 ctgggagagc agatgcgtca agacttcgac ctgttgaaca agtccatcct gcagaatatc
      601 aagatcttca gctcctccga gcgtcgtgtg ctgcacactg ctcagtactg gacccgtgct
      661 ctgttcggtg ctgacgagct gggttccgac gagatctcta tccgcaagga cctgctggac
      721 gactccaacg ctgctaagga cctcatggac aaagtcaaga agaagttgaa gcccctgctg
      781 cgcgagggca aagaggctcc tcctcaattc gcttggccct ccaagatgcc cgagccttac
      841 ctggtcatca agcgtgtggt cgagctgatg aactaccaca agaaaatcat ggataacaac
      901 ttcgccaaga aagacgtgaa ctccatgcag acccgttggt gcacctctga ggaccccagc
      961 ctgttcaaag agcgttggga caagcttttc aaagagttca acaacgccga gaaggtcgac
     1021 ccttccaaga tctccgagct gtacgacacc atgaagtacg acgctctgca caaccgtcag
     1081 ttcttggaga acatcttcga ccccggcctg cctaacgagg ctatcgctga cgaactcggt
     1141 tctcactccc tggtggatcg ttaccccatc aacgtcctgg ccaagaacaa cttcaagatc
     1201 atcgactccc actccatgaa caactccggc aagaactcct ccaactccgt gggttctctc
     1261 ggctgggtgc tcgagtctgg aaagacctca accgctcgta accccaagtc ctctagccag
     1321 ttcgacgagc cccgtttcat gcagctgcgc gaactgtaca agctggctaa ggtcctgttc
     1381 gacttcatct gccccaaaga gtacggtatc tccgacgctg agaagctgga catcggcctg
     1441 ctgacttctc tgcctctggc taagcagatc ctgaacgaca tcggcgacat gaagaaccgc
     1501 gagactcccg cttgtgtggc ttacttcacc aaagagtccc acatctacac cctgctcaac
     1561 atcatctacg aaagcggtat ccccatgagg atcgctcgca acgctctgcc cgaactggac
     1621 tacttgtccc agatcacctt cgagttgtac gagtccaccg acgcttccgg ccagaagtcc
     1681 cactctatcc gtctgaagat gtctcccggt tgccacactc aggaccctct ggacgtgcaa
     1741 ttggacgacc gtcactacat ctcttgcatc cctaagatct ccctgaccaa gcacctcgac
     1801 atggactacg tgcagcagaa gctgcgtaac aagttcaccc gtgtgatcat gcctccaaag
     1861 ttcactcccg tgaacatcac ctcataagcg gccgcactcg agtaggatcg aatcggactt
     1921 tcgaatctag agcctgcagt ctcgacaagc ttgtcgagaa gtactagagg atcataatca
     1981 gccataccac atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga
     2041 acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg
     2101 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     2161 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggatc tgatcactgc
     2221 ttgagcctag aagatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc
     2281 accgctgagc aataactatc ataaccccta gggtataccc atctaattgg aaccagataa
     2341 gtgaaatcta gttccaaact attttgtcat ttttaatttt cgtattagct tacgacgcta
     2401 cacccagttc ccatctattt tgtcactctt ccctaaataa tccttaaaaa ctccatttcc
     2461 acccctccca gttcccaact attttgtccg cccacaaccg gttgacttgg gtcaactgtc
     2521 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     2581 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     2641 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     2701 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     2761 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     2821 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     2881 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     2941 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     3001 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     3061 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     3121 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     3181 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     3241 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     3301 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattg
     3361 acttgggtcg ctcttcctgt ggatgcgcag atgccctgcg taagcgggtg tgggcggaca
     3421 ataaagtctt aaactgaaca aaatagatct aaactatgac aataaagtct taaactagac
     3481 agaatagttg taaactgaaa tcagtccagt tatgctgtga aaaagcatac tggacttttg
     3541 ttatggctaa agcaaactct tcattttctg aagtgcaaat tgcccgtcgt attaaagagg
     3601 ggcgtggcca agggcatgta aagactatat tcgcggcgtt gtgacaattt accgaacaac
     3661 tccgcggccg ggaagccgat ctcggcttga acgaattgtt aggtggcggt acttgggtcg
     3721 atatcaaagt gcatcacttc ttcccgtatg cccaactttg tatagagagc cactgcggga
     3781 tcgtcaccgt aatctgcttg cacgtagatc acataagcac caagcgcgtt ggcctcatgc
     3841 ttgaggagat tgatgagcgc ggtggcaatg ccctgcctcc ggtgctcgcc ggagactgcg
     3901 agatcataga tatagatctc actacgcggc tgctcaaact tgggcagaac gtaagccgcg
     3961 agagcgccaa caaccgcttc ttggtcgaag gcagcaagcg cgatgaatgt cttactacgg
     4021 agcaagttcc cgaggtaatc ggagtccggc tgatgttggg agtaggtggc tacgtctccg
     4081 aactcacgac cgaaaagatc aagagcagcc cgcatggatt tgacttggtc agggccgagc
     4141 ctacatgtgc gaatgatgcc catacttgag ccacctaact ttgttttagg gcgactgccc
     4201 tgctgcgtaa catcgttgct gctgcgtaac atcgttgctg ctccataaca tcaaacatcg
     4261 acccacggcg taacgcgctt gctgcttgga tgcccgaggc atagactgta caaaaaaaca
     4321 gtcataacaa gccatgaaaa ccgccactgc gccgttacca ccgctgcgtt cggtcaaggt
     4381 tctggaccag ttgcgtgagc gcatacgcta cttgcattac agtttacgaa ccgaacaggc
     4441 ttatgtcaac tgggttcgtg ccttcatccg tttccacggt gtgcgtcacc cggcaacctt
     4501 gggcagcagc gaagtcgcca taacttcgta tagcatacat tatacgaagt tatctgtaac
     4561 tataacggtc ctaaggtagc gagtttaaac actagtatcg attcgcgacc tactccggaa
     4621 ggtttaaacg tacccgtagt ggctatggca gggcttgccg ccccgacgtt ggctgcgagc
     4681 cctgggcctt cacccgaact tgggggttgg ggtggggaaa aggaagaaac gcgggcgtat
     4741 tggtcccaat ggggtctcgg tggggtatcg acagagtgcc agccctggga ccgaaccccg
     4801 cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta ttgccgtcat
     4861 agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc cccatctccc
     4921 ggtaccgcat gctccgattc gatcctactt gtacagctcg tccatgccga gagtgatccc
     4981 ggcggcggtc acgaactcca gcaggaccat gtgatcgcgc ttctcgttgg ggtctttgct
     5041 cagggcggac tggtagctca ggtagtggtt gtcgggcagc agcacggggc cgtcgccgat
     5101 gggggtgttc tgctggtagt ggtcggcgag ctgcacgctg ccgtcctcga tgttgtggcg
     5161 gatcttgaag ttcaccttga tgccgttctt ctgcttgtcg gccatgatat agacgttgtg
     5221 gctgttgtag ttgtactcca gcttgtgccc caggatgttg ccgtcctcct tgaagtcgat
     5281 acccttcagc tcgatgcggt tcaccagggt gtcgccctcg aacttcacct cggcgcgggt
     5341 cttgtagttg ccgtcgtcct tgaagaagat ggtgcgctcc tggacgtagc cttcgggcat
     5401 ggcggacttg aagaagtcgt gctgcttcat gtggtcgggg tagcgggcga agcactgcag
     5461 gccgtagccg aaggtggtca cgagggtggg ccagggcacg ggcagcttgc cggtggtgca
     5521 gatgaacttc agggtcagct tgccgtaggt ggcatcgccc tcgccctcgc cggacacgct
     5581 gaacttgtgg ccgtttacgt cgccgtccag ctcgaccagg atgggcacca ccccggtgaa
     5641 cagctcctcg cccttgctca ccatcatggt ttcggaccgc ccgggtgatc aagtcttcgt
     5701 cgagtgattg taaataaaat gtaatttaca gtatagtatt ttaattaata tacaaatgat
     5761 ttgataataa ttcttattta actataatat attgtgttgg gttgaattaa aggtccgtat
     5821 actagtatcg attcgcgacc tactccggaa tattaataga tcatggagat aattaaaatg
     5881 ataaccatct cgcaaataaa taagtatttt actgttttcg taacagtttt gtaataaaaa
     5941 aacctataaa tattccggat tattcatacc gtcccaccat cgggcgcgg
//
