LOCUS       Exported                7911 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Lentiviral vector to express H2B-iRFP670 under PGK promoter.
ACCESSION   .
VERSION     .
KEYWORDS    pLentiPGK DEST H2B-iRFP670
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7911)
  AUTHORS   Kudo T, Jeknic S, Macklin DN, Akhter S, Hughey JJ, Regot S, Covert 
            MW
  TITLE     Live-cell measurements of kinase activity in single cells using 
            translocation reporters.
  JOURNAL   Nat Protoc. 2018 Jan;13(1):155-169. doi: 10.1038/nprot.2017.128. 
            Epub 2017 Dec 21.
  PUBMED    29266096
REFERENCE   2  (bases 1 to 7911)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7911
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(46..65)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        237..616
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        618..816
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     771..791
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             834..1014
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1061..1186
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1683..1916
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2101..2145
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2443..2560
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     protein_bind    2631..2655
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    2684..2736
                     /label=MCS
                     /note="multiple cloning site"
     promoter        2758..3268
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3187..3205
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3356..3733
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     CDS             3788..4723
                     /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"
     protein_bind    complement(4747..4771)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    4799..5387
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4852..4872)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5270..5281)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5390..5406)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5391..5407)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5916..6096
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6158..6746)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6238..6257)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6917..7777)
                     /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     7540..7559
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7778..7882)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga
       61 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
      121 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
      181 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
      241 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      301 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      361 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      421 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      481 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      541 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      601 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      661 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      721 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      781 cggtaggcgt gtacggtggg aggtctatat aagcagcgcg ttttgcctgt actgggtctc
      841 tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta
      901 agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact
      961 ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtggcg
     1021 cccgaacagg gacttgaaag cgaaagggaa accagaggag ctctctcgac gcaggactcg
     1081 gcttgctgaa gcgcgcacgg caagaggcga ggggcggcga ctggtgagta cgccaaaaat
     1141 tttgactagc ggaggctaga aggagagaga tgggtgcgag agcgtcagta ttaagcgggg
     1201 gagaattaga tcgcgatggg aaaaaattcg gttaaggcca gggggaaaga aaaaatataa
     1261 attaaaacat atagtatggg caagcaggga gctagaacga ttcgcagtta atcctggcct
     1321 gttagaaaca tcagaaggct gtagacaaat actgggacag ctacaaccat cccttcagac
     1381 aggatcagaa gaacttagat cattatataa tacagtagca accctctatt gtgtgcatca
     1441 aaggatagag ataaaagaca ccaaggaagc tttagacaag atagaggaag agcaaaacaa
     1501 aagtaagacc accgcacagc aagcggccgg ccgctgatct tcagacctgg aggaggagat
     1561 atgagggaca attggagaag tgaattatat aaatataaag tagtaaaaat tgaaccatta
     1621 ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag agcagtggga
     1681 ataggagctt tgttccttgg gttcttggga gcagcaggaa gcactatggg cgcagcgtca
     1741 atgacgctga cggtacaggc cagacaatta ttgtctggta tagtgcagca gcagaacaat
     1801 ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac tcacagtctg gggcatcaag
     1861 cagctccagg caagaatcct ggctgtggaa agatacctaa aggatcaaca gctcctgggg
     1921 atttggggtt gctctggaaa actcatttgc accactgctg tgccttggaa tgctagttgg
     1981 agtaataaat ctctggaaca gatttggaat cacacgacct ggatggagtg ggacagagaa
     2041 attaacaatt acacaagctt aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa
     2101 aagaatgaac aagaattatt ggaattagat aaatgggcaa gtttgtggaa ttggtttaac
     2161 ataacaaatt ggctgtggta tataaaatta ttcataatga tagtaggagg cttggtaggt
     2221 ttaagaatag tttttgctgt actttctata gtgaatagag ttaggcaggg atattcacca
     2281 ttatcgtttc agacccacct cccaaccccg aggggacccg acaggcccga aggaatagaa
     2341 gaagaaggtg gagagagaga cagagacaga tccattcgat tagtgaacgg atcggcactg
     2401 cgtgcgccaa ttctgcagac aaatggcagt attcatccac aattttaaaa gaaaaggggg
     2461 gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac
     2521 taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag
     2581 cagagatcca gtttggttag taccgggccg gtggaattct gcagatatca acaagtttgt
     2641 acaaaaaagc aggctttaaa ggaaccaatt cagtcgactg gatctcgagc tcaagcttcg
     2701 aattctgcag tcgacggtac cgcgggcccg ggatccaccg gccggtcgcc accccacggg
     2761 gttggggttg cgccttttcc aaggcagccc tgggtttgcg cagggacgcg gctgctctgg
     2821 gcgtggttcc gggaaacgca gcggcgccga ccctgggtct cgcacattct tcacgtccgt
     2881 tcgcagcgtc acccggatct tcgccgctac ccttgtgggc cccccggcga cgcttcctgc
     2941 tccgccccta agtcgggaag gttccttgcg gttcgcggcg tgccggacgt gacaaacgga
     3001 agccgcacgt ctcactagta ccctcgcaga cggacagcgc cagggagcaa tggcagcgcg
     3061 ccgaccgcga tgggctgtgg ccaatagcgg ctgctcagca gggcgcgccg agagcagcgg
     3121 ccgggaaggg gcggtgcggg aggcggggtg tggggcggta gtgtgggccc tgttcctgcc
     3181 cgcgcggtgt tccgcattct gcaagcctcc ggagcgcacg tcggcagtcg gctccctcgt
     3241 tgaccgaatc accgacctct ctccccaggg ggatccaccg gtcgtcgaca gctcaagctt
     3301 cgaattctgc agtcgacggt accgcgggcc cgggatccac cggccggtcg ccaccatgcc
     3361 agagccagcg aagtctgctc ccgccccgaa aaagggctcc aagaaggcgg tgactaaggc
     3421 gcagaagaaa ggcggcaaga agcgcaagcg cagccgcaag gagagctatt ccatctatgt
     3481 gtacaaggtt ctgaagcagg tccaccctga caccggcatt tcgtccaagg ccatgggcat
     3541 catgaattcg tttgtgaacg acattttcga gcgcatcgca ggtgaggctt cccgcctggc
     3601 gcattacaac aagcgctcga ccatcacctc cagggagatc cagacggccg tgcgcctgct
     3661 gctgcctggg gagttggcca agcacgccgt gtccgagggt actaaggcca tcaccaagta
     3721 caccagcgct aagaacatgg ggcatggcac cggcagcacc ggcagcggca gctcatccgg
     3781 aactagtatg gcgcgtaagg tcgatctcac ctcctgcgat cgcgagccga tccacatccc
     3841 cggcagcatt cagccgtgcg gctgcctgct agcctgcgac gcgcaggcgg tgcggatcac
     3901 gcgcattacg gaaaatgccg gcgcgttctt tggacgcgaa actccgcggg tcggtgagct
     3961 actcgccgat tacttcggcg agaccgaagc ccatgcgctg cgcaacgcac tggcgcagtc
     4021 ctccgatcca aagcgaccgg cgctgatctt cggttggcgc gacggcctga ccggccgcac
     4081 cttcgacatc tcactgcatc gccatgacgg tacatcgatc atcgagttcg agcctgcggc
     4141 ggccgaacag gccgacaatc cgctgcggct gacgcggcag atcatcgcgc gcaccaaaga
     4201 actgaagtcg ctcgaagaga tggccgcacg ggtgccgcgc tatctgcagg cgatgctcgg
     4261 ctatcaccgc gtgatgttgt accgcttcgc ggacgacggc tccgggatgg tgatcggcga
     4321 ggcgaagcgc agcgacctcg agagctttct cggtcagcac tttccggcgt cgctggtccc
     4381 gcagcaggcg cggctactgt acttgaagaa cgcgatccgc gtggtctcgg attcgcgcgg
     4441 catcagcagc cggatcgtgc ccgagcacga cgcctccggc gccgcgctcg atctgtcgtt
     4501 cgcgcacctg cgcagcatct cgccctgcca tctcgaattt ctgcggaaca tgggcgtcag
     4561 cgcctcgatg tcgctgtcga tcatcattga cggcacgcta tggggattga tcatctgtca
     4621 tcattacgag ccgcgtgccg tgccgatggc gcagcgcgtc gcggccgaaa tgttcgccga
     4681 cttcttatcg ctgcacttca ccgccgccca ccaccaacgc taagcggccg cactcgagat
     4741 atctagaccc agctttcttg tacaaagtgg ttgatatcca gcacagtggc ggcccgataa
     4801 tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc
     4861 ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat
     4921 ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg
     4981 gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg
     5041 ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat
     5101 tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt
     5161 gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc
     5221 ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa
     5281 tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg
     5341 ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc gataccgtcg
     5401 acctcgatcg agacctagaa aaacatggag caatcacaag tagcaataca gcagctacca
     5461 atgctgattg tgcctggcta gaagcacaag aggaggagga ggtgggtttt ccagtcacac
     5521 ctcaggtacc tttaagacca atgacttaca aggcagctgt agatcttagc cactttttaa
     5581 aagaaaaggg gggactggaa gggctaattc actcccaacg aagacaagat atccttgatc
     5641 tgtggatcta ccacacacaa ggctacttcc ctgattggca gaactacaca ccagggccag
     5701 ggatcagata tccactgacc tttggatggt gctacaagct agtaccagtt gagcaagaga
     5761 aggtagaaga agccaatgaa ggagagaaca cccgcttgtt acaccctgtg agcctgcatg
     5821 ggatggatga cccggagaga gaagtattag agtggaggtt tgacagccgc ctagcatttc
     5881 atcacatggc ccgagagctg catccggact gtactgggtc tctctggtta gaccagatct
     5941 gagcctggga gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc
     6001 cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc
     6061 tcagaccctt ttagtcagtg tggaaaatct ctagcagcat gtgagcaaaa ggccagcaaa
     6121 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     6181 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     6241 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     6301 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     6361 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     6421 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     6481 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     6541 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     6601 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     6661 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     6721 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     6781 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     6841 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     6901 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     6961 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     7021 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     7081 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     7141 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     7201 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     7261 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     7321 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     7381 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     7441 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     7501 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     7561 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     7621 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     7681 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     7741 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     7801 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     7861 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac g
//
