LOCUS Exported 8405 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION mammalian expression of human Alpha7 (CHRNA7) with mGFP fused into M3-M4 loop. ACCESSION . VERSION . KEYWORDS pcDNA3.1-CHRNA7-mGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8405) AUTHORS Wang Y, Xiao C, Indersmitten T, Freedman R, Leonard S, Lester HA TITLE The duplicated alpha7 subunits assemble and form functional nicotinic receptors with the full-length alpha7. JOURNAL J Biol Chem. 2014 Sep 19;289(38):26451-63. doi: 10.1074/jbc.M114.582858. Epub 2014 Jul 23. PUBMED 25056953 REFERENCE 2 (bases 1 to 8405) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8405 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 15..394 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 395..598 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 549..569 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 643..662 /label=T7 /note="T7 promoter, forward primer" promoter 643..661 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 2034..2750 /codon_start=1 /product="enhanced GFP with monomerizing A206K mutation (Zacharias et al., 2002)" /label=mEGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(2079..2100) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(2340..2359) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 2687..2708 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(3607..3624) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 3613..3837 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 3883..4311 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3970..3989) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4180..4201 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(4320..4340) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 4325..4654 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 4505..4640 /label=SV40 ori /note="SV40 origin of replication" primer_bind 4567..4586 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 4703..5728 /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 5858..5979 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5895..5914) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 5949..5968 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(6028..6044) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(6028..6044) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(6041..6063) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 6052..6068 /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(6076..6106) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 6121..6142 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(6259..6276) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(6430..7018) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(6510..6529) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(7189..8049) /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 7812..7831 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(8050..8154) /gene="bla" /label=AmpR promoter primer_bind complement(8229..8248) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" ORIGIN 1 cagatatacg cgttgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc 61 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc 121 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt 181 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca 241 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg 301 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca 361 gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa 421 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa 481 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc 541 cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct 601 ctggctaact agagaaccca ctgcttactg gcttatcgaa attaatacga ctcactatag 661 ggagacccaa gctggctagc gtttaaactt aagcttggta ccgagctcgg atccactagt 721 aacggccgcc agtgtgctgg aattctgcag atatccatca cactggcggc cgctcgagcg 781 gactcaacat gcgctgctcg ccgggaggcg tctggctggc gctggccgcg tcgctcctgc 841 acgtgtccct gcaaggcgag ttccagagga agctttacaa ggagctggtc aagaactaca 901 atcccctgga gaggcccgtg gccaatgact cgcaaccact caccgtctac ttctccctga 961 gcctcctgca gatcatggac gtggatgaga agaaccaagt tttaaccacc aacatttggc 1021 tgcaaatgtc ttggacagat cactatttac agtggaatgt gtcagaatat ccaggggtga 1081 agactgttcg tttcccagat ggccagattt ggaaaccaga cattcttctc tataacagtg 1141 ctgatgagcg ctttgacgcc acattccaca ctaacgtgtt ggtgaattct tctgggcatt 1201 gccagtacct gcctccaggc atattcaaga gttcctgcta catcgatgta cgctggtttc 1261 cctttgatgt gcagcactgc aaactgaagt ttgggtcctg gtcttacgga ggctggtcct 1321 tggatctgca gatgcaggag gcagatatca gtggctatat ccccaatgga gaatgggacc 1381 tagtgggaat ccccggcaag aggagtgaaa ggttctatga gtgctgcaaa gagccctacc 1441 ctgatgtcac cttcacagtg accatgcgcc gcaggacact ctactatggc ctcaacctgc 1501 tgatcccctg tgtgctcatc tccgccctcg ccctgctggt gttcctgctt cctgcagatt 1561 ccggggagaa gatttccctg gggataacag tcttactctc tcttaccgtc ttcatgctgc 1621 tcgtggctga gatcatgccc gcaacatccg attcggtacc attgatagcc cagtacttcg 1681 ccagcaccat gatcatcgtg ggcctctcgg tggtggtgac ggtgatcgtg ctgcagtacc 1741 accaccacga ccccgacggg ggcaagatgc ccaagtggac cagagtcatc cttctgaact 1801 ggtgcgcgtg gttcctgcga atgaagaggc ccggggagga caaggtgcgc ccggcctgcc 1861 agcacaagca gcggcgctgc agcctggcca gtgtggagat gagcgccgtg gcgccgccgc 1921 ccgccagcaa cgggaacctg ctgtacatcg gcttccgcgg cctggacggc gtgcactgtg 1981 tcccgacccc cgactctggg gtagtgtgtg gccgcatggc ctgcgctggc gccatggtga 2041 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg 2101 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc 2161 tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga 2221 ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg 2281 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg 2341 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc 2401 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg 2461 agtacaacta caacagccac aacgtctata tcatggccga caagcagaag aacggcatca 2521 aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact 2581 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga 2641 gcacccagtc caagcttagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg 2701 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag gcaggcgcct 2761 cccccacgca cgatgagcac ctcctgcacg gtgggcaacc ccccgagggg gacccggact 2821 tggccaagat cctggaggag gtccgctaca ttgccaaccg cttccgctgc caggacgaaa 2881 gcgaggcggt ctgcagcgag tggaagttcg ccgcctgtgt ggtggaccgc ctgtgcctca 2941 tggccttctc ggtcttcacc atcatctgca ccatcggcat cctgatgtcg gctcccaact 3001 tcgtggaggc cgtgtccaaa gactttgcgt aaccacgcct ggttctgtac atgtggaaaa 3061 ctcacagatg ggcaaggcct ttggcttggc gagatttggg ggtgctaatc caggacagca 3121 ttacacgcca caactccagt gttcccttct ggctgtcagt cgtgttgctt acggtttctt 3181 tgttacttta ggtagtagaa tctcagcact ttgtttcata ttctcagatg ggctgataga 3241 tatccttggc acatccgtac catcggtcag cagggccact gagtagtcat tttgcccatt 3301 agcccactgc ctggaaagcc cttcggagag ctccccatgg ctcctcacca ccgagacagt 3361 tggttttgca tgtctgcatg aaggtctacc tgaaaattca acatttgctt tttgcttgtg 3421 tacaaaccca gattgaagct aaaataaacc agactcacta aatcctttcc aaaaaaaaaa 3481 aaaaaaaaaa aaccggaatt cctgcagccc gggggatcca ctagtccagt gtggtggaat 3541 tctgcagata tccagcacag tggcggccgc tcgagtctag agggcccgtt taaacccgct 3601 gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc 3661 cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg 3721 catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca 3781 agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt 3841 ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg 3901 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc 3961 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc 4021 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg 4081 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg 4141 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg 4201 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt 4261 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg 4321 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca 4381 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg 4441 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc 4501 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat 4561 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg 4621 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat 4681 tttcggatct gatcagcacg tgatgaaaaa gcctgaactc accgcgacgt ctgtcgagaa 4741 gtttctgatc gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga 4801 atctcgtgct ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg 4861 cgccgatggt ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc 4921 gattccggaa gtgcttgaca ttggggaatt cagcgagagc ctgacctatt gcatctcccg 4981 ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca 5041 gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt 5101 cggcccattc ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc 5161 gattgctgat ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc 5221 cgtcgcgcag gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca 5281 cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc 5341 ggtcattgac tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt 5401 cttctggagg ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca 5461 tccggagctt gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca 5521 actctatcag agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg 5581 cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag 5641 cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc 5701 cagcactcgt ccgagggcaa aggaatagca cgtgctacga gatttcgatt ccaccgccgc 5761 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca 5821 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa 5881 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca 5941 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac 6001 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc 6061 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta 6121 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa 6181 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat 6241 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg 6301 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc 6361 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt 6421 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag 6481 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc 6541 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc 6601 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt 6661 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt 6721 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc 6781 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa 6841 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa 6901 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg 6961 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga 7021 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg 7081 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg 7141 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt 7201 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact 7261 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat 7321 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg 7381 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg 7441 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat 7501 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc 7561 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt 7621 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc 7681 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga 7741 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc 7801 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa 7861 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta 7921 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg 7981 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg 8041 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat 8101 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt 8161 tccccgaaaa gtgccacctg acgtcgacgg atcgggagat ctcccgatcc cctatggtgc 8221 actctcagta caatctgctc tgatgccgca tagttaagcc agtatctgct ccctgcttgt 8281 gtgttggagg tcgctgagta gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg 8341 accgacaatt gcatgaagaa tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta 8401 cgggc //