LOCUS       sequence-214288-        5572 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  human BK beta3 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 5572)
  TITLE     BK channel plasmids
REFERENCE   2  (bases 1 to 5572)
  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 beta3 EGFP
FEATURES             Location/Qualifiers
     source          1..5572
                     /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"
     primer_bind     674..690
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     674..690
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     regulatory      1512..1521
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1518..1520,1521..1523,1524..2237)
                     /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(1518..1520,1521..1523,1524..2234)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1563..1584)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1824..1843)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2171..2192
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2360..2481
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2397..2416)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2451..2470
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2488..2943)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2625..2646)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2837..2856
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2970..3074
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3076..3433
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3076..3267
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3099..3119)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3284..3419
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3468..4262
                     /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             3468..4262
                     /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(3522..3541)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4132..4151
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4450..4469)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4494..4541
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4870..5458
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5359..5378
                     /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 caaaagctgg agctccaccg
      661 cggtggcggc cgctctagaa ctagtggatc ccccgggctg caggaattcc cttttctctc
      721 cactgccaat gacagccttt cctgcctcag ggaagaagag agagacagac tacagtgatg
      781 gagacccact agatgtgcac aagaggctgc catccagtac tggagaggac cgagccgtga
      841 tgctggggtt tgccatgatg ggcttctcag tcctaatgtt cttcttgctc ggaacaacca
      901 ttctaaagcc ttttatgctc agcattcaga gagaagaatc gacctgcact gccatccaca
      961 cagatatcat ggacgactgg ctggactgtg ccttcacctg tggtgtgcac tgccacggtc
     1021 aggggaagta cccatgtctt caggtgtttg tgaacctcag ccatccaggt cagaaagctc
     1081 tcctacatta taatgaagag gctgtccaga taaatcccaa gtgcttttac acacctaagt
     1141 gccaccaaga tagaaatgat ttgctcaaca gtgctctgga cataaaagaa ttcttcgatc
     1201 acaaaaatgg aactcccttt tcatgcttct acagtccagc cagccaatct gaagatgtca
     1261 ttcttataaa aaagtatgac caaatggcta tcttccactg tttattttgg ccttcactga
     1321 ctctgctagg tggtgccctg attgttggca tggtgagatt aacacaacac ctgtccttac
     1381 tgtgtgaaaa atatagcact gtagtcagag atgaggtagg tggaaaagta ccttatatag
     1441 aacagcatca gttcaaactg tgcattatga ggaggagcaa aggaagagca gagaaatcag
     1501 atccaccggt cgccaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca
     1561 tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg
     1621 agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc
     1681 ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct
     1741 accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc
     1801 aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt
     1861 tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg
     1921 gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg
     1981 ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg
     2041 gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc
     2101 tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga
     2161 agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg
     2221 acgagctgta caagtaaagc ggccgcgact ctagatcata atcagccata ccacatttgt
     2281 agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat
     2341 gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa
     2401 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     2461 caaactcatc aatgtatctt aaggcgtaaa ttgtaagcgt taatattttg ttaaaattcg
     2521 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
     2581 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
     2641 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     2701 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
     2761 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     2821 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     2881 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     2941 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     3001 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     3061 gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg
     3121 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     3181 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     3241 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     3301 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     3361 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     3421 aggcttttgc aaagatcgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     3481 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     3541 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     3601 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag
     3661 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     3721 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     3781 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     3841 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     3901 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     3961 tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc gaggatctcg
     4021 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     4081 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     4141 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     4201 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     4261 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
     4321 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     4381 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accctagggg
     4441 gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat
     4501 aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc
     4561 ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt
     4621 ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa
     4681 cgtcggggcg gcaggccctg ccatagcctc aggttactca tatatacttt agattgattt
     4741 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     4801 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     4861 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     4921 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     4981 aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg
     5041 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     5101 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     5161 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     5221 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     5281 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     5341 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     5401 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     5461 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
     5521 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgccatgc at
//
