LOCUS       Exported                7356 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Mammalian expression of SLC24A2.
ACCESSION   .
VERSION     .
KEYWORDS    Human NCKX2-AS-long-FlagTag (pcDNA3.1+)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7356)
  AUTHORS   Visser F, Valsecchi V, Annunziato L, Lytton J
  TITLE     Exchangers NCKX2, NCKX3, and NCKX4: identification of Thr-551 as a 
            key residue in defining the apparent K(+) affinity of NCKX2.
  JOURNAL   J Biol Chem. 2007 Feb 16;282(7):4453-62. Epub 2006 Dec 15.
  PUBMED    17172467
REFERENCE   2  (bases 1 to 7356)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7356
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1187..1210
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2946..2963)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2952..3176
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3222..3650
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3309..3328)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3519..3540
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3659..3679)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3664..3994
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3845..3980
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3907..3926
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4061..4855
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4115..4134)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4725..4744
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5029..5150
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5066..5085)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5120..5139
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5199..5215)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5199..5215)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5212..5234)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5223..5239
                     /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(5247..5277)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5292..5313
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5430..5447)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5601..6189)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5681..5700)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6360..7220)
                     /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     6983..7002
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7221..7325)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccatggatct gcaacaaagc accaccatca cttccctaga
      961 gaaatggtgt ttggatgagt cactgtctgg ctgcagaaga cattatagtg tcaagaaaaa
     1021 actgaagtta attcgagtct taggcctttt catgggtctg gtagccatta gcactgtctc
     1081 attttcaatc agtgcctttt ctgagacaga tacacagagc acaggagagg ccagtgttgt
     1141 aagtggccct agggtagcac agggttacca tcagagaact ctcttagatt acaaggacga
     1201 cgatgacaag tatactccac agccacctct ttctaaggaa ggcgagtctg agaatagtac
     1261 agatcacgcc caaggagact acccgaaaga catcttttcc cttgaggaga gaagaaaagg
     1321 tgcgatcatt ctgcatgtca ttggaatgat ctacatgttc atagccttag ccattgtctg
     1381 tgatgagttc tttgttcctt ctttgactgt catcactgaa aaactgggca tctctgatga
     1441 tgtggctgga gccaccttca tggctgcagg agggtcagcc ccagaacttt tcacatctct
     1501 cataggggta tttatcgctc acagcaacgt tggcataggc acaattgtag gttcagcagt
     1561 attcaacatc ctctttgtta ttggcatgtg tgctctgttt tctagagaaa tcttaaacct
     1621 gacatggtgg ccgctctttc gagatgtgtc tttctacatt gttgacttga tcatgctgat
     1681 catatttttc ctggataatg tcatcatgtg gtgggaaagc ttgcttctct taacagctta
     1741 tttttgctat gtggttttca tgaaattcaa cgtccaagta gaaaaatggg tgaagcaaat
     1801 gataaaccgc aataaggtcg tcaaggtgac agcaccagaa gcccaagcaa agccatctgc
     1861 agccagggac aaggatgaac cgactctacc ggctaagccg cgtctccagc gaggtggaag
     1921 ctctgcctcc ctccacaaca gtctcatgag gaatagcatc ttccaactca tgatacacac
     1981 ccttgaccca ctcgccgaag aacttggatc atatggaaaa ctaaaatatt atgacacaat
     2041 gactgaagaa gggaggttca gagaaaaggc ttcaattctc cacaagatcg ccaagaagaa
     2101 atgtcatgtg gatgagaacg agcggcagaa tggggctgcc aaccacgtgg aaaaaattga
     2161 gcttccaaac agcaccagca cagatgttga aatgacacca tccagtgatg cttcagaacc
     2221 tgtacaaaat ggaaatctct cccacaacat tgaaggtgca gaagcccaga ccgctgatga
     2281 ggaggaggac cagcctctca gccttgcctg gccttctgaa acccgcaagc aagtcacgtt
     2341 tctgattgtt ttccccatag tgtttcctct ctggattacg ttacctgacg ttcgcaaacc
     2401 ttcatcgagg aagttttttc ccatcacgtt ctttggctcc attacctgga ttgcagtatt
     2461 ctcttacttg atggtctggt gggcgcacca ggttggagag acaattggca tcagtgaaga
     2521 gattatgggc ctgaccatct tggctgctgg gacctccatc cctgatctta tcaccagtgt
     2581 catagtggcc cggaaggggt taggggacat ggctgtgtcc agctctgttg gaagcaacat
     2641 ttttgacatc actgtagggc tcccactgcc ctggctcctg tacaccgtca ttcacagatt
     2701 ccagccagtg gctgtcagca gcaatggcct tttctgtgcc atcgtccttc tcttcatcat
     2761 gctgctcttc gtcatcctct ctatcgccct ctgcaagtgg cgaatgaaca aaatcctggg
     2821 cttcatcatg tttggcctct actttgtgtt cctggtggtg agcgttctcc tagaagacag
     2881 aattcttaca tgccccgtct ccatctagct cgagtctaga gggcccgttt aaacccgctg
     2941 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     3001 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     3061 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     3121 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     3181 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     3241 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     3301 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     3361 tcaagctcta aatcggggca tccctttagg gttccgattt agtgctttac ggcacctcga
     3421 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     3481 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     3541 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tggggatttc
     3601 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     3661 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc caggcaggca gaagtatgca
     3721 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     3781 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     3841 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     3901 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg
     3961 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat
     4021 tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt
     4081 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     4141 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     4201 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct
     4261 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     4321 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     4381 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     4441 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     4501 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     4561 agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac
     4621 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     4681 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     4741 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     4801 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     4861 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat
     4921 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     4981 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt
     5041 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     5101 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt
     5161 ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga
     5221 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     5281 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     5341 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     5401 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     5461 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     5521 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     5581 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     5641 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     5701 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     5761 gcctttctcc cttcgggaag cgtggcgctt tctcaatgct cacgctgtag gtatctcagt
     5821 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     5881 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     5941 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     6001 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     6061 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     6121 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     6181 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     6241 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     6301 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     6361 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     6421 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     6481 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     6541 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     6601 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     6661 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     6721 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     6781 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     6841 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     6901 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     6961 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     7021 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     7081 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     7141 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     7201 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     7261 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     7321 ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
