LOCUS       sequence-214287-        5716 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 5716)
  TITLE     BK channel plasmids
REFERENCE   2  (bases 1 to 5716)
  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..5716
                     /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      1656..1665
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1662..1664,1665..1667,1668..2381)
                     /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(1662..1664,1665..1667,1668..2378)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1707..1728)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1968..1987)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2315..2336
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2504..2625
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2541..2560)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2595..2614
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2632..3087)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2769..2790)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2981..3000
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3114..3218
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3220..3577
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3220..3411
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3243..3263)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3428..3563
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3612..4406
                     /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             3612..4406
                     /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(3666..3685)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4276..4295
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4594..4613)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4638..4685
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5014..5602
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5503..5522
                     /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 taatcctatc caagtatgca gtacgctctt gggtcgtctc atgagaccca
      721 ggggcatgtt ggaaagaact gagagaaaga gcaacaaagc ggcgagtggt gtgagagggc
      781 agcacgcgct gtggggccct tccagagaaa tgtactgaaa aagtctacgc aatgtctggg
      841 atttgctaaa caatacctgg aaagcagaca ggtttttttg ccattcctcc aggacatcca
      901 ccataaggaa aggagaccct ggaccaacat tctctaagat gtttatatgg accagtggcc
      961 ggacctcttc atcttataga catgatgaaa agagaaatat ttaccagaaa atcagggacc
     1021 atgacctcct ggacaaaagg aaaacagtca cagcactgaa ggcaggagag gaccgagcta
     1081 ttctcctggg actggctatg atggtgtgct ccatcatgat gtattttctg ctgggaatca
     1141 cactcctgcg ctcatacatg cagagcgtgt ggaccgaaga gtctcaatgc accttgctga
     1201 atgcgtccat cacggaaaca tttaactgct ccttcagctg tggtccagac tgctggaaac
     1261 tttctcagta cccctgcctc caggtgtacg ttaacctgac ttcttccggg gaaaagctcc
     1321 tcctctacca cacagaagag acaataaaaa tcaatcagaa gtgctcctat atacctaaat
     1381 gtggaaaaaa ttttgaagaa tccatgtccc tggtgaatgt tgtcatggaa aacttcagga
     1441 agtatcaaca cttctcctgc tattctgacc cagaaggaaa ccagaagagt gttatcctaa
     1501 ccaaactcta cagttccaac gtgctgttcc attcactctt ctggccaacc tgtatgatgg
     1561 ctgggggtgt ggcaattgtt gccatggtga aacttacaca gtacctctcc ctactatgtg
     1621 agaggatcca acggatcaat agagatccac cggtcgccac catggtgagc aagggcgagg
     1681 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
     1741 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
     1801 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct
     1861 acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
     1921 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     1981 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     2041 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     2101 acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca
     2161 agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca
     2221 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg
     2281 ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     2341 ccgccgggat cactctcggc atggacgagc tgtacaagta aagcggccgc gactctagat
     2401 cataatcagc cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct
     2461 ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc
     2521 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     2581 actgcattct agttgtggtt tgtccaaact catcaatgta tcttaaggcg taaattgtaa
     2641 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     2701 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     2761 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     2821 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     2881 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     2941 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     3001 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     3061 cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa
     3121 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     3181 cctgataaat gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa agaaccagct
     3241 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     3301 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
     3361 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     3421 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     3481 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     3541 gtgaggaggc ttttttggag gcctaggctt ttgcaaagat cgatcaagag acaggatgag
     3601 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     3661 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     3721 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     3781 tgaatgaact gcaagacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     3841 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     3901 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     3961 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     4021 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     4081 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcga
     4141 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     4201 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     4261 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     4321 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     4381 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     4441 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     4501 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     4561 ggagttcttc gcccacccta gggggaggct aactgaaaca cggaaggaga caataccgga
     4621 aggaacccgc gctatgacgg caataaaaag acagaataaa acgcacggtg ttgggtcgtt
     4681 tgttcataaa cgcggggttc ggtcccaggg ctggcactct gtcgataccc caccgagacc
     4741 ccattggggc caatacgccc gcgtttcttc cttttcccca ccccaccccc caagttcggg
     4801 tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag cctcaggtta
     4861 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     4921 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     4981 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     5041 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     5101 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     5161 ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     5221 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     5281 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     5341 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     5401 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     5461 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     5521 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     5581 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     5641 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     5701 tattaccgcc atgcat
//
