LOCUS       sequence-491518-        4560 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  Expression plasmid for iRFP670 driven from the EF1a promoter. 
            Co-transfection marker for transfection experiments..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4560)
  AUTHORS   Kabaria SR, Bae Y, Ehmann ME, Lende-Dorn BA, Beitz AM, Peterman EL, 
            Love KS, Ploessl DS, Galloway KE
  TITLE     Programmable promoter editing for precise control of transgene 
            expression.
  JOURNAL   Nat Biotechnol. 2025 Oct 13. doi: 10.1038/s41587-025-02854-y.
  PUBMED    41083901
REFERENCE   2  (bases 1 to 4560)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 4, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Biotechnol. 2025 Oct 13. doi: 10.1038/s41587-025-02854-y."
COMMENT     Sequence Label: EF1a-iRFP670-SV40_pKG726
FEATURES             Location/Qualifiers
     source          1..4560
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    10..109
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        161..1338
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          392..1330
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1287..1307
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             1343..2278
                     /codon_start=1
                     /product="near-infrared fluorescent protein with an 
                     emission peak at 670 nm, engineered from a bacterial 
                     phytochrome (Shcherbakova and Verkhusha, 2013)"
                     /label=iRFP670
                     /note="derived from Rhodopseudomonas palustris RpBphP6"
                     /translation="MARKVDLTSCDREPIHIPGSIQPCGCLLACDAQAVRITRITENAG
                     AFFGRETPRVGELLADYFGETEAHALRNALAQSSDPKRPALIFGWRDGLTGRTFDISLH
                     RHDGTSIIEFEPAAAEQADNPLRLTRQIIARTKELKSLEEMAARVPRYLQAMLGYHRVM
                     LYRFADDGSGMVIGEAKRSDLESFLGQHFPASLVPQQARLLYLKNAIRVVSDSRGISSR
                     IVPEHDASGAALDLSFAHLRSISPCHLEFLRNMGVSASMSLSIIIDGTLWGLIICHHYE
                     PRAVPMAQRVAAEMFADFLSLHFTAAHHQR"
     polyA_signal    2288..2409
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2325..2344)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2379..2398
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(2441..2540)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2652..2671)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2663..3472
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPGDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2730..2749)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3562..4150
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4051..4070
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      4315..4401
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4493..4520
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     4528..4551
                     /label=pENTR-F
                     /note="pENTR vectors, forward primer"
ORIGIN
        1 ctcgggcccc aaataatgat tttattttga ctgatagtga cctgttcgtt gcaacaaatt
       61 gataagcaat gcttttttat aatgccaact ttgtacaaaa aagcaggctc caccatggga
      121 accaattcag tcgactggat ccggtacagg aatccggtac ggctccggtg cccgtcagtg
      181 ggcagagcgc acatcgccca cagtccccga gaagttgtgg ggaggggtcg gcaattgaac
      241 cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg
      301 cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct
      361 ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt cccgcgggcc
      421 tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct ggctgcagta
      481 cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga ggccttgcgc
      541 ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct ggggccgccg
      601 cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt ctctagccat
      661 ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc ttgtaaatgc
      721 gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg acggggcccg
      781 tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac cgagaatcgg
      841 acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc cgccgtgtat
      901 cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag cggaaagatg
      961 gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct cgggagagcg
     1021 ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg tcgcttcatg
     1081 tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga gcttttggag
     1141 tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc acactgagtg
     1201 ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg aatttgccct
     1261 ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa gtttttttct
     1321 tccatttcag gtgtcgtgct aaatggcgcg taaggtcgat ctcacctcct gcgatcgcga
     1381 gccgatccac atccccggca gcattcagcc gtgcggctgc ctgctagcct gcgacgcgca
     1441 ggcggtgcgg atcacgcgca ttacggaaaa tgccggcgcg ttctttggac gcgaaactcc
     1501 gcgggtcggt gagctactcg ccgattactt cggcgagacc gaagcccatg cgctgcgcaa
     1561 cgcactggcg cagtcctccg atccaaagcg accggcgctg atcttcggtt ggcgcgacgg
     1621 cctgaccggc cgcaccttcg acatctcact gcatcgccat gacggtacat cgatcatcga
     1681 gttcgagcct gcggcggccg aacaggccga caatccgctg cggctgacgc ggcagatcat
     1741 cgcgcgcacc aaagaactga agtcgctcga agagatggcc gcacgggtgc cgcgctatct
     1801 gcaggcgatg ctcggctatc accgcgtgat gttgtaccgc ttcgcggacg acggctccgg
     1861 gatggtgatc ggcgaggcga agcgcagcga cctcgagagc tttctcggtc agcactttcc
     1921 ggcgtcgctg gtcccgcagc aggcgcggct actgtacttg aagaacgcga tccgcgtggt
     1981 ctcggattcg cgcggcatca gcagccggat cgtgcccgag cacgacgcct ccggcgccgc
     2041 gctcgatctg tcgttcgcgc acctgcgcag catctcgccc tgccatctcg aatttctgcg
     2101 gaacatgggc gtcagcgcct cgatgtcgct gtcgatcatc attgacggca cgctatgggg
     2161 attgatcatc tgtcatcatt acgagccgcg tgccgtgccg atggcgcagc gcgtcgcggc
     2221 cgaaatgttc gccgacttct tatcgctgca cttcaccgcc gcccaccacc aacgctaaag
     2281 aacaaccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     2341 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     2401 tgtatcttat tagtaatgaa ccggtccatc ggatatctag acccagcttt cttgtacaaa
     2461 gttggcatta taagaaagca ttgcttatca atttgttgca acgaacaggt cactatcagt
     2521 caaaataaaa tcattatttg ccatccagct gcagctctgg cccgtgtctc aaaatctctg
     2581 atgttacatt gcacaagata aaaatatatc atcatgaaca ataaaactgt ctgcttacat
     2641 aaacagtaat acaaggggtg ttatgagcca tattcaacgg gaaacgtcga ggccgcgatt
     2701 aaattccaac atggatgctg atttatatgg gtataaatgg gctcgcgata atgtcgggca
     2761 atcaggtgcg acaatctatc gcttgtatgg gaagcccgat gcgccagagt tgtttctgaa
     2821 acatggcaaa ggtagcgttg ccaatgatgt tacagatgag atggtcagac taaactggct
     2881 gacggaattt atgcctcttc cgaccatcaa gcattttatc cgtactcctg gtgatgcatg
     2941 gttactcacc actgcgatcc ccggaaaaac agcattccag gtattagaag aatatcctga
     3001 ttcaggtgaa aatattgttg atgcgctggc agtgttcctg cgccggttgc attcgattcc
     3061 tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt ctagctcagg cgcaatcacg
     3121 aatgaataac ggtttggttg atgcgagtga ttttgatgac gagcgtaatg gctggcctgt
     3181 tgaacaagtc tggaaagaaa tgcataaact tttgccattc tcaccggatt cagtcgtcac
     3241 tcatggtgat ttctcacttg ataaccttat ttttgacgag gggaaattaa taggttgtat
     3301 tgatgttgga cgagtcggaa tcgcagaccg ataccaggat cttgccatcc tatggaactg
     3361 cctcggtgag ttttctcctt cattacagaa acggcttttt caaaaatatg gtattgataa
     3421 tcctgatatg aataaattgc agtttcattt gatgctcgat gagtttttct aatcagaatt
     3481 ggttaattgg ttgtaacatt attcagattg ggccccgttc cactgagcgt cagaccccgt
     3541 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     3601 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     3661 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     3721 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     3781 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     3841 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     3901 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     3961 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     4021 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     4081 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     4141 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     4201 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgctag
     4261 catggatctc ggggacgtct aactactaag cgagagtagg gaactgccag gcatcaaata
     4321 aaacgaaagg ctcagtcgga agactgggcc tttcgtttta tctgttgttt gtcggtgaac
     4381 gctctcctga gtaggacaaa tccgccggga gcggatttga acgttgtgaa gcaacggccc
     4441 ggagggtggc gggcaggacg cccgccataa actgccaggc atcaaactaa gcagaaggcc
     4501 atcctgacgg atggcctttt tgcgtttcta caaactcttc ctgttagtta gttacttaag
//
