LOCUS       sequence-459549-        8955 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express ALFANb-CAAX.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8955)
  AUTHORS   Madern MF, Yang S, Witteveen O, Segeren HA, Bauer M, Tanenbaum ME
  TITLE     Long-term imaging of individual ribosomes reveals ribosome 
            cooperativity in mRNA translation.
  JOURNAL   Cell. 2025 Jan 23:S0092-8674(25)00045-5. doi: 
            10.1016/j.cell.2025.01.016.
  PUBMED    39892379
REFERENCE   2  (bases 1 to 8955)
  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: "Cell. 2025 
            Jan 23:S0092-8674(25)00045-5. doi: 10.1016/j.cell.2025.01.016."
COMMENT     Sequence Label: pHR-ALFANb-GB1-CAAX-WPRE
FEATURES             Location/Qualifiers
     source          1..8955
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(16..34)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(17..34)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(324..343)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(557..576)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     676..698
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(736..754)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        822..926
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(927..995,996..1787)
                     /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     complement(1145..1164)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1958..2546
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2447..2466
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2700..2717
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(2787..2807)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2792..3121
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2972..3107
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          4255..4320
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             4450..4470
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    4895..5029
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4932..4951)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4986..5005
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     LTR             5197..5830
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    5877..6002
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6499..6732
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6917..6961
                     /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    7246..7363
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        7516..7923
                     /label=SFFV promoter
                     /note="spleen focus-forming virus long terminal repeat 
                     (LTR) promoter"
     regulatory      7957..7966
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             8338..8499
                     /codon_start=1
                     /product="B1 domain of Streptococcal protein G   "
                     /label=GB1
                     /note="effective as a solubilizing fusion partner (Cheng 
                     and Patel, 2004)"
                     /translation="YKLILNGKTLKGETTTEAVDAATAEKVFKQYANDNGVDGEWTYDD
                     ATKTFTVTE"
     LTR             8713..8946
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
ORIGIN
        1 attaattccg tgtattctat agtgtcacct aaatcgtatg tgtatgatac ataaggttat
       61 gtattaattg tagccgcgtt ctaacgacaa tatgtacaag cctaattgtg tagcatctgg
      121 cttactgaag cagaccctat catctctctc gtaaactgcc gtcagagtcg gtttggttgg
      181 acgaaccttc tgagtttctg gtaacgccgt cccgcacccg gaaatggtca gcgaaccaat
      241 cagcagggtc atcgctagcc agatcctcta cgccggacgc atcgtggccg gcatcaccgg
      301 cgccacaggt gcggttgctg gcgcctatat cgccgacatc accgatgggg aagatcgggc
      361 tcgccacttc gggctcatga gcgcttgttt cggcgtgggt atggtggcag gccccgtggc
      421 cgggggactg ttgggcgcca tctccttgca tgcaccattc cttgcggcgg cggtgctcaa
      481 cggcctcaac ctactactgg gctgcttcct aatgcaggag tcgcataagg gagagcgtcg
      541 aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc
      601 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
      661 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
      721 gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
      781 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
      841 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
      901 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
      961 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     1021 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     1081 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     1141 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     1201 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     1261 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     1321 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     1381 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     1441 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     1501 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     1561 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     1621 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     1681 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     1741 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     1801 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     1861 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     1921 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     1981 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     2041 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     2101 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     2161 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     2221 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     2281 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     2341 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     2401 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     2461 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     2521 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     2581 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     2641 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     2701 gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc
     2761 gttggccgat tcattaatgc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     2821 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     2881 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     2941 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     3001 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     3061 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     3121 aaagcttgga cacaagacag gcttgcgaga tatgtttgag aataccactt tatcccgcgt
     3181 cagggagagg cagtgcgtaa aaagacgcgg actcatgtga aatactggtt tttagtgcgc
     3241 cagatctcta taatctcgcg caacctattt tcccctcgaa cactttttaa gccgtagata
     3301 aacaggctgg gacacttcac atgagcgaaa aatacatcgt cacctgggac atgttgcaga
     3361 tccatgcacg taaactcgca agccgactga tgccttctga acaatggaaa ggcattattg
     3421 ccgtaagccg tggcggtctg taccgggtgc gttactggcg cgtgaactgg gtattcgtca
     3481 tgtcgatacc gtttgtattt ccagctacga tcacgacaac cagcgcgagc ttaaagtgct
     3541 gaaacgcgca gaaggcgatg gcgaaggctt catcgttatt gatgacctgg tggataccgg
     3601 tggtactgcg gttgcgattc gtgaaatgta tccaaaagcg cactttgtca ccatcttcgc
     3661 aaaaccggct ggtcgtccgc tggttgatga ctatgttgtt gatatcccgc aagatccctg
     3721 gattgaacag ccgtgggata tgggcgtcgt attcgtcccg ccaatctccg gtcgctaatc
     3781 ttttcaacgc ctggcactgc cgggcgttgt tctttttaac ttcaggcggg ttacaatagt
     3841 ttccagtaag tattctggag gctgcatcca tgacacaggc aaacctgagc gaaaccctgt
     3901 tcaaaccccg ctttaaacat cctgaaacct cgacgctagt ccgccgcttt aatcacggcg
     3961 cacaaccgcc tgtgcagtcg gcccttgatg gtaaaaccat ccctcactgg tatcgcatga
     4021 ttaaccgtct gatgtggatc tggcgcggca ttgacccacg cgaaatcctc gacgtccagg
     4081 cacgtattgt gatgagcgat gccgaacgta ccgacgatga tttatacgat acggtgattg
     4141 gctaccgtgg cggcaactgg atttatgagt gggccccgga tctttgtgaa ggaaccttac
     4201 ttctgtggtg tgacataatt ggacaaacta cctacagaga tttaaagctc taaggtaaat
     4261 ataaaatttt taagtgtata atgtgttaaa ctactgattc taattgtttg tgtattttag
     4321 attccaacct atggaactga tgaatgggag cagtggtgga atgcctttaa tgaggaaaac
     4381 ctgttttgct cagaagaaat gccatctagt gatgatgagg ctactgctga ctctcaacat
     4441 tctactcctc caaaaaagaa gagaaaggta gaagacccca aggactttcc ttcagaattg
     4501 ctaagttttt tgagtcatgc tgtgtttagt aatagaactc ttgcttgctt tgctatttac
     4561 accacaaagg aaaaagctgc actgctatac aagaaaatta tggaaaaata ttctgtaacc
     4621 tttataagta ggcataacag ttataatcat aacatactgt tttttcttac tccacacagg
     4681 catagagtgt ctgctattaa taactatgct caaaaattgt gtacctttag ctttttaatt
     4741 tgtaaagggg ttaataagga atatttgatg tatagtgcct tgactagaga tcataatcag
     4801 ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa
     4861 cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg
     4921 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     4981 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggatca actggataac
     5041 tcaagctaac caaaatcatc ccaaacttcc caccccatac cctattacca ctgccaatta
     5101 cctgtggttt catttactct aaacctgtga ttcctctgaa ttattttcat tttaaagaaa
     5161 ttgtatttgt taaatatgta ctacaaactt agtagttgga agggctaatt cactcccaaa
     5221 gaagacaaga tatccttgat ctgtggatct accacacaca aggctacttc cctgattagc
     5281 agaactacac accagggcca ggggtcagat atccactgac ctttggatgg tgctacaagc
     5341 tagtaccagt tgagccagat aaggtagaag aggccaataa aggagagaac accagcttgt
     5401 tacaccctgt gagcctgcat gggatggatg acccggagag agaagtgtta gagtggaggt
     5461 ttgacagccg cctagcattt catcacgtgg cccgagagct gcatccggag tacttcaaga
     5521 actgctgata tcgagcttgc tacaagggac tttccgctgg ggactttcca gggaggcgtg
     5581 gcctgggcgg gactggggag tggcgagccc tcagatcctg catataagca gctgcttttt
     5641 gcctgtactg ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta
     5701 gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc
     5761 cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa
     5821 atctctagca gtggcgcccg aacagggact tgaaagcgaa agggaaacca gaggagctct
     5881 ctcgacgcag gactcggctt gctgaagcgc gcacggcaag aggcgagggg cggcgactgg
     5941 tgagtacgcc aaaaattttg actagcggag gctagaagga gagagatggg tgcgagagcg
     6001 tcagtattaa gcgggggaga attagatcgc gatgggaaaa aattcggtta aggccagggg
     6061 gaaagaaaaa atataaatta aaacatatag tatgggcaag cagggagcta gaacgattcg
     6121 cagttaatcc tggcctgtta gaaacatcag aaggctgtag acaaatactg ggacagctac
     6181 aaccatccct tcagacagga tcagaagaac ttagatcatt atataataca gtagcaaccc
     6241 tctattgtgt gcatcaaagg atagagataa aagacaccaa ggaagcttta gacaagatag
     6301 aggaagagca aaacaaaagt aagaccaccg cacagcaagc ggccggccgc tgatcttcag
     6361 acctggagga ggagatatga gggacaattg gagaagtgaa ttatataaat ataaagtagt
     6421 aaaaattgaa ccattaggag tagcacccac caaggcaaag agaagagtgg tgcagagaga
     6481 aaaaagagca gtgggaatag gagctttgtt ccttgggttc ttgggagcag caggaagcac
     6541 tatgggcgca gcgtcaatga cgctgacggt acaggccaga caattattgt ctggtatagt
     6601 gcagcagcag aacaatttgc tgagggctat tgaggcgcaa cagcatctgt tgcaactcac
     6661 agtctggggc atcaagcagc tccaggcaag aatcctggct gtggaaagat acctaaagga
     6721 tcaacagctc ctggggattt ggggttgctc tggaaaactc atttgcacca ctgctgtgcc
     6781 ttggaatgct agttggagta ataaatctct ggaacagatt tggaatcaca cgacctggat
     6841 ggagtgggac agagaaatta acaattacac aagcttaata cactccttaa ttgaagaatc
     6901 gcaaaaccag caagaaaaga atgaacaaga attattggaa ttagataaat gggcaagttt
     6961 gtggaattgg tttaacataa caaattggct gtggtatata aaattattca taatgatagt
     7021 aggaggcttg gtaggtttaa gaatagtttt tgctgtactt tctatagtga atagagttag
     7081 gcagggatat tcaccattat cgtttcagac ccacctccca accccgaggg gacccgacag
     7141 gcccgaagga atagaagaag aaggtggaga gagagacaga gacagatcca ttcgattagt
     7201 gaacggatct cgacggtatc gccaaatggc agtattcatc cacaatttta aaagaaaagg
     7261 ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca
     7321 aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga
     7381 cagcagagat ccagtttgga tcgataagct tgatatcgaa ttcctgcagc cccgataaaa
     7441 taaaagattt tatttagtct ccagaaaaag gggggaatga aagaccccac ctgtaggttt
     7501 ggcaagctag ctgcagtaac gccattttgc aaggcatgga aaaataccaa accaagaata
     7561 gagaagttca gatcaagggc gggtacatga aaatagctaa cgttgggcca aacaggatat
     7621 ctgcggtgag cagtttcggc cccggcccgg ggccaagaac agatggtcac cgcagtttcg
     7681 gccccggccc gaggccaaga acagatggtc cccagatatg gcccaaccct cagcagtttc
     7741 ttaagaccca tcagatgttt ccaggctccc ccaaggacct gaaatgaccc tgcgccttat
     7801 ttgaattaac caatcagcct gcttctcgct tctgttcgcg cgcttctgct tcccgagctc
     7861 tataaaagag ctcacaaccc ctcactcggc gcgccagtcc tccgacagac tgagtcgccc
     7921 gggggggatc tggagctctc gagaattctc acgcgtgcca ccatggaggt gcagctgcag
     7981 gagagtggag gtggactggt gcagcccgga ggtagtctgc gactgagctg caccgccagc
     8041 ggcgtgacca tcagcgccct gaacgccatg gccatgggct ggtaccgcca ggcccccggc
     8101 gagcgccgcg tgatggtggc cgccgtgagc gagcgcggca acgccatgta ccgcgagagc
     8161 gtgcagggcc gcttcaccgt gacccgcgac ttcaccaaca agatggtgag cctgcagatg
     8221 gacaacctga agcccgagga caccgccgtg tactactgcc acgtgctgga ggaccgcgtg
     8281 gacagcttcc acgactactg gggccagggc acccaggtga ccgtgagcag cttggagtac
     8341 aagcttatcc tgaacggtaa aaccctgaaa ggtgaaacca ccaccgaagc tgttgacgct
     8401 gctaccgcgg aaaaagtttt caaacagtac gctaacgaca acggtgttga cggtgaatgg
     8461 acctacgacg acgctaccaa aaccttcacg gtaaccgaag gatccggtgg aggttctggt
     8521 ggtggatctg gtggaggttc tggaggtgga aaaatgtcca aggatggtaa gaaaaagaag
     8581 aagaagtcaa aaaccaagtg tgttatcatg taaactagtt aagcggccgc gactctagag
     8641 tcgacccttt aagaccaatg acttacaagg cagctgtaga tcttagccac tttttaaaag
     8701 aaaagggggg actggaaggg ctaattcact cccaacgaag acaagatctg ctttttgctt
     8761 gtactgggtc tctctggtta gaccagatct gagcctggga gctctctggc taactaggga
     8821 acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc
     8881 tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct
     8941 ctagcagcat ctaga
//
