LOCUS       sequence-498273-        5806 bp    DNA     circular SYN 26-JAN-2026
DEFINITION  In vitro transcription (SP6 promotor) or mammalian expression (CMV 
            promotor) of a flourenscent reporter containing a box-b hairpin 
            without two editible stop codons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5806)
  TITLE     Cifuentes Lab Plasmids 2025
REFERENCE   2  (bases 1 to 5806)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 26, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pCS2_RFP_2xRecEnhancedEdited_1xBoxB_EYFP
FEATURES             Location/Qualifiers
     source          1..5806
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        678..696
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     678..695
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      737..746
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             746..769
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1499..1552
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     protein_bind    1618..1636
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     CDS             1637..1693
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1709..1735
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(1736..1738,1739..1741,1742..2455)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1781..1802)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2042..2061)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2389..2410
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2501..2635
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2525..2544)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2579..2598
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2748..2768)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2748..2766)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2787..2803)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2787..2803)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2800..2822)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2811..2827
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(2835..2841,2842..2859,2860..2865))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2880..2901
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3018..3035)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3189..3777)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3269..3288)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3948..4739,4740..4808))
                     /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     4571..4590
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4809..4913)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4939..5394)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5076..5097)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5288..5307
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        join(5465..5806,1..643)
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cctatgccaa tatggctatt ggccaggttc aatactatgt attggcccta tgccatatag
       61 tattccatat atgggttttc ctattgacgt agatagcccc tcccaatggg cggtcccata
      121 taccatatat ggggcttcct aataccgccc atagccactc ccccattgac gtcaatggtc
      181 tctatatatg gtctttccta ttgacgtcat atgggcggtc ctattgacgt atatggcgcc
      241 tcccccattg acgtcaatta cggtaaatgg cccgcctggc tcaatgccca ttgacgtcaa
      301 taggaccacc caccattgac gtcaatggga tggctcattg cccattcata tccgttctca
      361 cgccccctat tgacgtcaat gacggtaaat ggcccacttg gcagtacatc aatatctatt
      421 aatagtaact tggcaagtac attactattg gaagtacgcc agggtacatt ggcagtactc
      481 ccattgacgt caatggcggt aaatggcccg cgatggctgc caagtacatc cccattgacg
      541 tcaatgggga ggggcaatga cgcaaatggg cgttccattg acgtaaatgg gcggtaggcg
      601 tgcctaatgg gaggtctata taagcaatgc tcgtttaggg aaccgccatt ctgcctgggg
      661 acgtcggagc aagcttgatt taggtgacac tatagaatac aagctacttg ttctttttgc
      721 aggatcccat cgattcgcca ccatggacta caaggacgac gacgacaagg tgagcaaagg
      781 agaggaactg attaaagaaa acatgcacat gaagctgtac atggagggaa cagtgaacaa
      841 ccaccacttc aagtgtacat ctgagggaga gggaaagcct tatgagggaa cacagacaat
      901 gagaatcaag gtggtggagg gaggacctct gccttttgct tttgacatcc tggctaccag
      961 cttcatgtat ggaagcagaa cattcatcaa ccacacacag ggaatccctg acttcttcaa
     1021 gcagagcttc cctgagggat tcacatggga gagagtgaca acatatgagg atggaggagt
     1081 gctgacagct acacaggaca caagcctcca ggatggatgt ctgatctaca atgtgaagat
     1141 cagaggagtg aacttcccta gcaatggacc tgtgatgcag aagaagacac tgggatggga
     1201 ggctaacaca gagatgctgt accctgctga tggaggactg gagggaagaa cagacatggc
     1261 tctgaagctg gtgggaggag gacacctgat ctgtaacttc aagacaacat acagaagcaa
     1321 gaagcctgct aagaacctga agatgcctgg agtgtactat gtggaccaca gactggagag
     1381 aatcaaggag gctgacaagg agacatatgt ggagcagcat gaggtggctg tggctagata
     1441 ctgtgacctg cctagcaagc tgggacacaa gctgaatgga atggatgagc tgtacaagga
     1501 gggcaggggg agcctcctaa cttgcggaga cgtcgaggag aatcctggcc cagatatcga
     1561 tccagatcca gatgacaaaa tctggatctg gatcgaaaat tctgagctcg gcacgacggg
     1621 ccctgaagaa gggcccgcca cgaacttctc tctgttaaag caagcaggag acgtggaaga
     1681 aaaccccggt cccggatccg ccaccatgta tccgtacgac gtaccggatt atgcgatggt
     1741 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     1801 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     1861 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     1921 gaccaccttc ggctacggcc tgcagtgctt cgcccgctac cccgaccaca tgaagcagca
     1981 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     2041 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     2101 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     2161 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
     2221 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     2281 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     2341 gagctaccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     2401 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtagcaagg
     2461 cctctcgagc ctctagaact atagtgagtc gtattacgta gatccagaca tgataagata
     2521 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga
     2581 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa
     2641 caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaatt
     2701 cgcggccgcg gcgccaatgc attgggcccg gtacccagct tttgttccct ttagtgaggg
     2761 ttaattgcgc gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     2821 ctcacaattc cacacaacat acgagccggg agcataaagt gtaaagcctg gggtgcctaa
     2881 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     2941 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     3001 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     3061 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     3121 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     3181 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     3241 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     3301 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     3361 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     3421 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     3481 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     3541 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     3601 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     3661 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     3721 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     3781 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     3841 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     3901 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     3961 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     4021 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     4081 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     4141 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     4201 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     4261 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     4321 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     4381 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     4441 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     4501 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     4561 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     4621 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     4681 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     4741 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     4801 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     4861 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     4921 ccccgaaaag tgccacctaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt
     4981 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
     5041 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
     5101 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     5161 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     5221 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     5281 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
     5341 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcccatt
     5401 cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac
     5461 gccagtcgac cgccaattca atatggcgta tatggactca tgccaattca atatggtgga
     5521 tctggacctg tgccaattca atatggcgta tatggactcg tgccaattca atatggtgga
     5581 tctggacccc agccaattca atatggcgga cttggcacca tgccaattca atatggcgga
     5641 cctggcactg tgccaactgg ggaggggtct acttggcacg gtgccaagtt tgaggagggg
     5701 tcttggccct gtgccaagtc cgccatattg aattggcatg gtgccaataa tggcggccat
     5761 attggctata tgccaggatc aatatatagg caatatccaa tatggc
//
