LOCUS       sequence-304471-        5985 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression construct for producing Avi tagged Methionyl-tRNA 
            synthetase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5985)
  TITLE     Graslund lab plasmids
REFERENCE   2  (bases 1 to 5985)
  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: MARS_pNIC-CTB10H
FEATURES             Location/Qualifiers
     source          1..5985
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     869..888
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(908..985)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1191..1210
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1294..1313
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1294..1312
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1313..1337
                     /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)."
     CDS             2059..2079
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2086..2130
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2131..2160
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     primer_bind     complement(2213..2231)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2227..2274
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2311..2766
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2398..2417)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2608..2629
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2858..3673)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2858..3673)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3581..3600
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3665..3684
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3795..4383
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4284..4303
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4537..4554
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4569..4708
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4634..4653)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4794..4816
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4810..5001)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5810..5985,1..907))
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
ORIGIN
        1 ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aacggcggga
       61 tataacatga gctgtcttcg gtatcgtcgt atcccactac cgagatatcc gcaccaacgc
      121 gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca
      181 gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca
      241 tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt
      301 tatgccagcc agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg
      361 cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat
      421 gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa
      481 cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga
      541 tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc
      601 cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa
      661 tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca
      721 gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg
      781 ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca
      841 cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg
      901 gtttcacatt caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa
      961 aggttttgcg ccattcgatg gtgtccggga tctcgacgct ctcccttatg cgactcctgc
     1021 attaggaagc agcccagtag taggttgagg ccgttgagca ccgccgccgc aaggaatggt
     1081 gcatgcaagg agatggcgcc caacagtccc ccggccacgg ggcctgccac catacccacg
     1141 ccgaaacaag cgctcatgag cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg
     1201 gcgatatagg cgccagcaac cgcacctgtg gcgccggtga tgccggccac gatgcgtccg
     1261 gcgtagagga tcgagatctc gatcccgcga aattaatacg actcactata ggggaattgt
     1321 gagcggataa caattcccct ctagaaataa ttttgtttaa ccttaagaag gagatatact
     1381 atgagactgt tcgtgagtga tggcgtcccg ggttgcttgc cggtgctggc cgccgccggg
     1441 agagcccggg gcagagcaga ggtgctcatc agcactgtag gcccggaaga ttgtgtggtc
     1501 ccgttcctga cccggcctaa ggtccctgtc ttgcagctgg atagcggcaa ctacctcttc
     1561 tccactagtg caatctgccg atattttttt ttgttatctg gctgggagca agatgacctc
     1621 actaaccagt ggctggaatg ggaagcgaca gagctgcagc cagctttgtc tgctgccctg
     1681 tactatttag tggtccaagg caagaagggg gaagatgttc ttggttcagt gcggagagcc
     1741 ctgactcaca ttgaccacag cttgagtcgt cagaactgtc ctttcctggc tggggagaca
     1801 gaatctctag ccgacattgt tttgtgggga gccctatacc cattactgca agatcccgcc
     1861 tacctccctg aggagctgag tgccctgcac agctggttcc agacactgag tacccaggaa
     1921 ccatgtcagc gagctgcaga gactgtactg aaacagcaag gtgtcctggc tctccggcct
     1981 tacctccaaa agcagcccca gcccagcccc gctgagggaa gggctgtcac caatgagcct
     2041 gaggaggagg agctggcaga gaacctctac ttccaatcgg gtggcggtct gaacgacatc
     2101 ttcgaggctc agaaaatcga atggcacgaa catcatcatc atcaccacca ccaccaccac
     2161 tgagatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag
     2221 caataactag cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa
     2281 ggaggaacta tatccggatt ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg
     2341 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     2401 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     2461 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     2521 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     2581 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     2641 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     2701 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta
     2761 caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     2821 aatacattca aatatgtatc cgctcatgaa ttaattctta gaaaaactca tcgagcatca
     2881 aatgaaactg caatttattc atatcaggat tatcaatacc atatttttga aaaagccgtt
     2941 tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga tcctggtatc
     3001 ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa
     3061 taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag aatggcaaaa
     3121 gtttatgcat ttctttccag acttgttcaa caggccagcc attacgctcg tcatcaaaat
     3181 cactcgcatc aaccaaaccg ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc
     3241 gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc aggaacactg
     3301 ccagcgcatc aacaatattt tcacctgaat caggatattc ttctaatacc tggaatgctg
     3361 ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg ataaaatgct
     3421 tgatggtcgg aagaggcata aattccgtca gccagtttag tctgaccatc tcatctgtaa
     3481 catcattggc aacgctacct ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc
     3541 catacaatcg atagattgtc gcacctgatt gcccgacatt atcgcgagcc catttatacc
     3601 catataaatc agcatccatg ttggaattta atcgcggcct agagcaagac gtttcccgtt
     3661 gaatatggct cataacaccc cttgtattac tgtttatgta agcagacagt tttattgttc
     3721 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     3781 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     3841 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     3901 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
     3961 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     4021 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     4081 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     4141 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     4201 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     4261 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     4321 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     4381 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     4441 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     4501 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     4561 gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcaat
     4621 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct
     4681 atcgctacgt gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc
     4741 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     4801 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag
     4861 ctcatcagcg tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg cgtccagctc
     4921 gttgagtttc tccagaagcg ttaatgtctg gcttctgata aagcgggcca tgttaagggc
     4981 ggttttttcc tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt
     5041 aatgataccg atgaaacgag agaggatgct cacgatacgg gttactgatg atgaacatgc
     5101 ccggttactg gaacgttgtg agggtaaaca actggcggta tggatgcggc gggaccagag
     5161 aaaaatcact cagggtcaat gccagcgctt cgttaataca gatgtaggtg ttccacaggg
     5221 tagccagcag catcctgcga tgcagatccg gaacataatg gtgcagggcg ctgacttccg
     5281 cgtttccaga ctttacgaaa cacggaaacc gaagaccatt catgttgttg ctcaggtcgc
     5341 agacgttttg cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta
     5401 accagtaagg caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg
     5461 cacccgtggg gccgccatgc cggcgataat ggcctgcttc tcgccgaaac gtttggtggc
     5521 gggaccagtg acgaaggctt gagcgagggc gtgcaagatt ccgaataccg caagcgacag
     5581 gccgatcatc gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc
     5641 cggcacctgt cctacgagtt gcatgataaa gaagacagtc ataagtgcgg cgacgatagt
     5701 catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg
     5761 agatcccggt gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc
     5821 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     5881 aggcggtttg cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca
     5941 gctgattgcc cttcaccgcc tggccctgag agagttgcag caagc
//
