LOCUS       sequence-218921-        5719 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  human BK beta2 subunit in C-term translational frame with EGFP 
            (EGFP-N1).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5719)
  TITLE     BK channel plasmids
REFERENCE   2  (bases 1 to 5719)
  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: BK beta2 EGFP
FEATURES             Location/Qualifiers
     source          1..5719
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        619..637
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      1659..1668
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1665..1667,1668..1670,1671..2384)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1665..1667,1668..1670,1671..2381)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1710..1731)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1971..1990)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2318..2339
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2507..2628
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2544..2563)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2598..2617
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2635..3090)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2772..2793)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2984..3003
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3117..3221
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3223..3580
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3223..3414
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3246..3266)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3431..3566
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3615..4409
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3615..4409
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3669..3688)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4279..4298
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4597..4616)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4641..4688
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5017..5605
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5506..5525
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagaa ttaaccctca ctaaagggaa taagcttgcg gccgccaact
      661 tttttttttt tcttaatcct atccaagtat gcagtacgct cttgggtcgt ctcatgagac
      721 ccaggggcat gttggaaaga actgagagaa agagcaacaa agcggcgagt ggtgtgagag
      781 ggcagcacgc gctgtggggc ccttccagag aaatgtactg aaaaagtcta cgcaatgtct
      841 gggatttgct aaacaatacc tggaaagcag acaggttttt ttgccattcc tccaggacat
      901 ccaccataag gaaaggagac cctggaccaa cattctctaa gatgtttata tggaccagtg
      961 gccggacctc ttcatcttat agacatgatg aaaaaagaaa tatttaccag aaaatcaggg
     1021 accatgacct cctggacaaa aggaaaacag tcacagcact gaaggcagga gaggaccgag
     1081 ctattctcct gggactggct atgatggtgt gctccatcat gatgtatttt ctgctgggaa
     1141 tcacactcct gcgctcatac atgcagagcg tgtggaccga agagtctcaa tgcaccttgc
     1201 tgaatgcgtc catcacggaa acatttaact gctccttcag ctgtggtcca gactgctgga
     1261 aactttctca gtacccctgc ctccaggtgt acgttaacct gacttcttcc ggggaaaagc
     1321 tcctcctcta ccacacagaa gagacaataa aaatcaatca gaagtgctcc tatataccta
     1381 aatgtggaaa aaattttgaa gaatccatgt ccctggtgaa tgttgtcatg gaaaacttca
     1441 ggaagtatca acacttctcc tgctattctg acccagaagg aaaccagaag agtgttatcc
     1501 taaccaaact ctacagttcc aacgtgctgt tccattcact cttctggcca acctgtatga
     1561 tggctggggg tgtggcaatt gttgccatgg tgaaacttac acagtacctc tccctactat
     1621 gtgagaggat ccaacggatc aatagagatc caccggtcgc caccatggtg agcaagggcg
     1681 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
     1741 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
     1801 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga
     1861 cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca
     1921 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     1981 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     2041 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact
     2101 acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact
     2161 tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga
     2221 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt
     2281 ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     2341 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaaagcggc cgcgactcta
     2401 gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca
     2461 cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc
     2521 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     2581 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttaag gcgtaaattg
     2641 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     2701 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     2761 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     2821 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     2881 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     2941 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     3001 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     3061 ccgcgcttaa tgcgccgcta cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg
     3121 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     3181 aaccctgata aatgcttcaa taatattgaa aaaggaagag tcctgaggcg gaaagaacca
     3241 gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag
     3301 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
     3361 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
     3421 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
     3481 actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa
     3541 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa gagacaggat
     3601 gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
     3661 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg
     3721 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg
     3781 ccctgaatga actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc
     3841 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg
     3901 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca
     3961 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc
     4021 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg
     4081 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg
     4141 cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata
     4201 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg
     4261 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat
     4321 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct
     4381 tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca
     4441 agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt
     4501 gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat
     4561 gctggagttc ttcgcccacc ctagggggag gctaactgaa acacggaagg agacaatacc
     4621 ggaaggaacc cgcgctatga cggcaataaa aagacagaat aaaacgcacg gtgttgggtc
     4681 gtttgttcat aaacgcgggg ttcggtccca gggctggcac tctgtcgata ccccaccgag
     4741 accccattgg ggccaatacg cccgcgtttc ttccttttcc ccaccccacc ccccaagttc
     4801 gggtgaaggc ccagggctcg cagccaacgt cggggcggca ggccctgcca tagcctcagg
     4861 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     4921 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     4981 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     5041 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     5101 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     5161 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     5221 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     5281 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     5341 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     5401 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     5461 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     5521 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     5581 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     5641 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     5701 ccgtattacc gccatgcat
//
