LOCUS       sequence-423961-        8477 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of the Blue Fluorescent Protein - P2A - full length 
            protein kinase R from chinook salmon.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8477)
  TITLE     Collet mammalian expression vectors
REFERENCE   2  (bases 1 to 8477)
  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"
COMMENT     Sequence Label: pcDNA3.1-Hyg-BFP-P2A-otPKR-FL
FEATURES             Location/Qualifiers
     source          1..8477
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(65..84)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        256..635
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        636..839
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     790..810
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     884..903
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        884..902
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1650..1706
                     /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             complement(2527..2538)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3920..3937)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3926..4150
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4196..4624
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4283..4302)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4493..4514
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4633..4653)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4638..4967
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4818..4953
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4880..4899
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5016..6041
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             5016..6041
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    6171..6292
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6208..6227)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6262..6281
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6341..6357)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6341..6357)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6354..6376)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6365..6381
                     /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(6389..6395,6396..6413,6414..6419))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6434..6455
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6572..6589)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6743..7331)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6823..6842)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7502..8293,8294..8362))
                     /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     8125..8144
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8363..8467)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc
       61 tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt
      121 tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg
      181 acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg
      241 ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca attacggggt
      301 cattagttca tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc
      361 ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag
      421 taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc
      481 acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg
      541 gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc
      601 agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca
      661 atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca
      721 atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg
      781 ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctc
      841 tctggctaac tagagaaccc actgcttact ggcttatcga aattaatacg actcactata
      901 gggagaccca agctggctag cgtttaaacg ggccctctag actcgagatg agcgagctga
      961 ttaaggagaa catgcacatg aagctgtaca tggagggcac cgtggacaac catcacttca
     1021 agtgcacatc cgagggcgaa ggcaagccct acgagggcac ccagaccatg agaatcaagg
     1081 tggtcgaggg cggccctctc cccttcgcct tcgacatcct ggctactagc ttcctctacg
     1141 gcagcaagac cttcatcaac cacacccagg gcatccccga cttcttcaag cagtccttcc
     1201 ctgagggctt cacatgggag agagtcacca catacgaaga cgggggcgtg ctgaccgcta
     1261 cccaggacac cagcctccag gacggctgcc tcatctacaa cgtcaagatc agaggggtga
     1321 acttcacatc caacggccct gtgatgcaga agaaaacact cggctgggag gccttcaccg
     1381 agacgctgta ccccgctgac ggcggcctgg aaggcagaaa cgacatggcc ctgaagctcg
     1441 tgggcgggag ccatctgatc gcaaacatca agaccacata tagatccaag aaacccgcta
     1501 agaacctcaa gatgcctggc gtctactatg tggactacag actggaaaga atcaaggagg
     1561 ccaacaacga gacctacgtc gagcagcacg aggtggcagt ggccagatac tgcgacctcc
     1621 ctagcaaact ggggcacgat ggaagcggag ctactaactt cagcctgctg aagcaggctg
     1681 gagacgtgga ggagaaccct ggacctatgg attccacaaa ctacatctcc atactgtgcg
     1741 agtatgctca gagacaacgt cagatctctg acatcaagtt tgaggaggtt ggaacagaag
     1801 gaccggatca cctgaaaacc ttcactctga gggtggtgat taaaggccat gcctatccta
     1861 acggggttgg aaagaacaag aaagaggcca aacagaatgc tgctaaacat gctctggctg
     1921 gcatgatgga gacgacagag cagaaggatg aacatgctct ggctggagtg gcggagacga
     1981 cagagcagaa ggatgcatcg gtttcatgtc ctccgtcacc ggctcagtct ccagcccctc
     2041 ctgtccgagc agttatctcc cagcccaact acgtgtgctg gctgaatgaa cacagccaga
     2101 agaacaagct gtctctgaag gctttggagg aaacgcgtgt tggacccaac aacacctcac
     2161 aatgctgtag gtatgtggta ggtgagaaag aatacccaga gggctttggg aacacgaaga
     2221 aggaggctaa ggaggaagca gcaatgcaga tctatctgga gttatgtggc cgtagtcaga
     2281 ctacagggac tgcagatgaa aactgcaatg gcaccactgg aaaacacaag gaggaattga
     2341 atccaaaggt tgtggtcctc agtgagaagg ttgaacacat gaacatgagc ccagaacaga
     2401 atgttccagc acgatacagc ttcattacaa cagaggagac caacttcata ggtatactca
     2461 accactactg tcagaaaacc aaacggtttc ctgactttaa actggtggag aagagcggac
     2521 cgtcacacga ccctcaattt gtttataaag tattgattga ccagagagaa tacccaaatg
     2581 gtttgggtaa gacagccaaa caggccaaac aacaagctgc tcagttagca tggtctgctc
     2641 ttcaagaaca gtctgactgg aacagccagg tgtcttgcag atcaacagtt tcagaagatg
     2701 gaggactatc ctctctaacc ccctccagta catgggaatc ccaggatgca acaccatcat
     2761 ccttgagtat tccaggaagt accagtgagt ccatcatatt cgcagactcc gttaacgtca
     2821 gctcccccaa ggttgaaagc cctgtaaatg tgaagcctaa aataagacta gcacctaact
     2881 tcctgatgtc accggtcaag accaaacagg aaggtcccga tttaaaaggc aacaatttag
     2941 gaaccttgtc aagtggaagg acatccaacc aaccaattaa atctaggttt ttatcagagt
     3001 ttgattcaat agagaagatt ggcaaaggag gctttggcaa cgtttataag gcgcgacggg
     3061 aactggagca gaagtatttt gccgtgaaga tagtactgag taaagggaaa gctaagcggg
     3121 aggttggtgc cttagcagat cttcagcacc ccaacatagt acgctactac accgcctggc
     3181 tggaggacac cgcgtaccga tgcgacacca actctgagag cgacaccact tcagattcag
     3241 gcagcagttc gtcctctgag ttcctgtaca tccagatgga actatgtgac aagaggactc
     3301 taaaagtgtg gatcgacgag aggaacgctc atcggaaacc caaacgcagg gaggagagtc
     3361 tacacatcac ccagcagata gtaaacgggg tagaatacat ccactccaag aaactcctac
     3421 acagagacct aaagcctgct aacatcatgt tcggtatgag tgacggagag gggaagggag
     3481 aggtaaagat tggggacttt ggtctggtga cggcagaaga taacgacaac gatgagaacc
     3541 tgctggagag aacgaaaaag acaggaacca agtcttacat ggcccccgaa cagaggaacc
     3601 agacctccta tgacagaaag gtggacatct ttgctttggg tctgatctac ttcgaactgc
     3661 tctggaacct gtctgggatg gagaaagcag aggtctggaa tgatgtcaga agccagagtt
     3721 ttcctcccca gttcaacacg cagttcaact tagagaacaa ggtgatagag tcgatgctgt
     3781 gtgccaaccc agaggataga ccagatgcca ggcagctgaa gataaaactg attgagtgtt
     3841 cttgtgttct gaccagagat caaaacttcc atcaagacaa cagaacagtc taaaagctta
     3901 agtttaaacc gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     3961 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     4021 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     4081 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg
     4141 ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg
     4201 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     4261 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     4321 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     4381 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     4441 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     4501 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     4561 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     4621 aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag
     4681 gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag
     4741 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc
     4801 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     4861 atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat
     4921 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag
     4981 cttgtatatc cattttcgga tctgatcagc acgtgatgaa aaagcctgaa ctcaccgcga
     5041 cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg atgcagctct
     5101 cggagggcga agaatctcgt gctttcagct tcgatgtagg agggcgtgga tatgtcctgc
     5161 gggtaaatag ctgcgccgat ggtttctaca aagatcgtta tgtttatcgg cactttgcat
     5221 cggccgcgct cccgattccg gaagtgcttg acattgggga attcagcgag agcctgacct
     5281 attgcatctc ccgccgtgca cagggtgtca cgttgcaaga cctgcctgaa accgaactgc
     5341 ccgctgttct gcagccggtc gcggaggcca tggatgcgat cgctgcggcc gatcttagcc
     5401 agacgagcgg gttcggccca ttcggaccgc aaggaatcgg tcaatacact acatggcgtg
     5461 atttcatatg cgcgattgct gatccccatg tgtatcactg gcaaactgtg atggacgaca
     5521 ccgtcagtgc gtccgtcgcg caggctctcg atgagctgat gctttgggcc gaggactgcc
     5581 ccgaagtccg gcacctcgtg cacgcggatt tcggctccaa caatgtcctg acggacaatg
     5641 gccgcataac agcggtcatt gactggagcg aggcgatgtt cggggattcc caatacgagg
     5701 tcgccaacat cttcttctgg aggccgtggt tggcttgtat ggagcagcag acgcgctact
     5761 tcgagcggag gcatccggag cttgcaggat cgccgcggct ccgggcgtat atgctccgca
     5821 ttggtcttga ccaactctat cagagcttgg ttgacggcaa tttcgatgat gcagcttggg
     5881 cgcagggtcg atgcgacgca atcgtccgat ccggagccgg gactgtcggg cgtacacaaa
     5941 tcgcccgcag aagcgcggcc gtctggaccg atggctgtgt agaagtactc gccgatagtg
     6001 gaaaccgacg ccccagcact cgtccgaggg caaaggaata gcacgtgcta cgagatttcg
     6061 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     6121 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     6181 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     6241 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     6301 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     6361 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     6421 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     6481 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     6541 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     6601 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     6661 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     6721 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     6781 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     6841 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     6901 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     6961 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     7021 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     7081 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     7141 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     7201 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     7261 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     7321 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     7381 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     7441 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     7501 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     7561 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     7621 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     7681 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     7741 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     7801 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     7861 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     7921 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     7981 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     8041 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     8101 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     8161 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     8221 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     8281 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     8341 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     8401 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     8461 ttccgcgcac atttccc
//
