LOCUS       sequence-304473-        7049 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression construct for producing Avi tagged Asparaginyl-tRNA 
            synthetase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7049)
  TITLE     Graslund lab plasmids
REFERENCE   2  (bases 1 to 7049)
  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: NARS_pNIC-Bio3
FEATURES             Location/Qualifiers
     source          1..7049
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     95..112
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    127..266
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(192..211)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     352..374
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(368..559)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1368..2450)
                     /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
                     EACKAAVHNLLAQRVSGLIINYQLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     2412..2431
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2451..2528)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     2734..2753
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2837..2856
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2837..2855
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2856..2880
                     /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)."
     RBS             2895..2917
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2928..2945
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2970..2990
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             4653..4697
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4750..4767
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4820..4838)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4834..4881
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4918..5373
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5005..5024)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5215..5236
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5465..6280)
                     /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(5465..6280)
                     /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     6188..6207
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6272..6291
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6402..6990
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6891..6910
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
       61 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga
      121 agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcaatgg
      181 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac actccgctat
      241 cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct
      301 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
      361 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct
      421 catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt
      481 tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg
      541 ttttttcctg tttggtcact gatgcctccg tgtaaggggg atttctgttc atgggggtaa
      601 tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat gaacatgccc
      661 ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa
      721 aaatcactca gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta
      781 gccagcagca tcctgcgatg cagatccgga acataatggt gcagggcgct gacttccgcg
      841 tttccagact ttacgaaaca cggaaaccga agaccattca tgttgttgct caggtcgcag
      901 acgttttgca gcagcagtcg cttcacgttc gctcgcgtat cggtgattca ttctgctaac
      961 cagtaaggca accccgccag cctagccggg tcctcaacga caggagcacg atcatgcgca
     1021 cccgtggggc cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg
     1081 gaccagtgac gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc
     1141 cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg
     1201 gcacctgtcc tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca
     1261 tgtcccgcgc ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgag
     1321 atcccggtgc ctaatgagtg agctaactta cattaattgc gttgcgctca ctgcccgctt
     1381 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     1441 gcggtttgcg tattgggcgc cagggtggtt tttcttttca ccagtgagac gggcaacagc
     1501 tgattgccct tcaccgcctg gccctgagag agttgcagca agcggtccac gctggtttgc
     1561 cccagcaggc gaaaatcctg tttgatggtg gttaacggcg ggatataaca tgagctgtct
     1621 tcggtatcgt cgtatcccac taccgagata tccgcaccaa cgcgcagccc ggactcggta
     1681 atggcgcgca ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc agtgggaacg
     1741 atgccctcat tcagcatttg catggtttgt tgaaaaccgg acatggcact ccagtcgcct
     1801 tcccgttccg ctatcggctg aatttgattg cgagtgagat atttatgcca gccagccaga
     1861 cgcagacgcg ccgagacaga acttaatggg cccgctaaca gcgcgatttg ctggtgaccc
     1921 aatgcgacca gatgctccac gcccagtcgc gtaccgtctt catgggagaa aataatactg
     1981 ttgatgggtg tctggtcaga gacatcaaga aataacgccg gaacattagt gcaggcagct
     2041 tccacagcaa tggcatcctg gtcatccagc tgatagttaa tgatcagccc actgacgcgt
     2101 tgcgcgagaa gattgtgcac cgccgcttta caggcttcga cgccgcttcg ttctaccatc
     2161 gacaccacca cgctggcacc cagttgatcg gcgcgagatt taatcgccgc gacaatttgc
     2221 gacggcgcgt gcagggccag actggaggtg gcaacgccaa tcagcaacga ctgtttgccc
     2281 gccagttgtt gtgccacgcg gttgggaatg taattcagct ccgccatcgc cgcttccact
     2341 ttttcccgcg ttttcgcaga aacgtggctg gcctggttca ccacgcggga aacggtctga
     2401 taagagacac cggcatactc tgcgacatcg tataacgtta ctggtttcac attcaccacc
     2461 ctgaattgac tctcttccgg gcgctatcat gccataccgc gaaaggtttt gcgccattcg
     2521 atggtgtccg ggatctcgac gctctccctt atgcgactcc tgcattagga agcagcccag
     2581 tagtaggttg aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca aggagatggc
     2641 gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac aagcgctcat
     2701 gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat aggcgccagc
     2761 aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga ggatcgagat
     2821 ctcgatcccg cgaaattaat acgactcact ataggggaat tgtgagcgga taacaattcc
     2881 cctctagaaa taattttgtt taactttaag aaggagatat acatatgcac catcatcatc
     2941 atcattcttc tggtgtagat ctgggtaccg agaacctgta cttccaatcc atggtgctag
     3001 cagagctgta cgtctctgac cgagagggaa gcgatgccac gggagatgga accaaggaga
     3061 aaccatttaa aacaggtcta aaggctttga tgacagtagg gaaagaacca tttcctacca
     3121 tttacgtaga ttcacaaaaa gaaaatgaga ggtggaatgt tatttctaaa tcacagttga
     3181 agaacattaa aaagatgtgg catagggaac aaatgaagag tgaatcccgg gaaaagaaag
     3241 aggcagaaga tagtttacga agagaaaaga acctggaaga agcaaagaag attaccatta
     3301 aaaatgatcc aagtctccca gagccaaaat gtgtgaagat tggtgcgtta gaaggatata
     3361 gaggccaaag agtaaaggtg tttggctggg tccacaggct gcgcaggcaa ggaaagaatt
     3421 taatgtttct ggtgttgcga gatggtacag gttatcttca gtgtgtcttg gcggatgagt
     3481 tgtgtcagtg ctacaatgga gttctcttgt ccacggagag cagtgttgca gtgtatggaa
     3541 tgctaaatct taccccaaag ggcaagcagg ctccaggtgg ccatgagctg agttgtgact
     3601 tctgggaact aattgggttg gcccctgctg gaggagctga caacctgatc aatgaggagt
     3661 ctgacgttga tgtccagctc aacaacagac acatgatgat ccgaggagaa aacatgtcca
     3721 aaatcctaaa agcacgatcc atggtcacca ggtgctttag agatcacttc tttgataggg
     3781 ggtactatga agttactcct ccaacattag tgcaaacaca agtagaaggt ggtgccacac
     3841 tcttcaagct tgactatttt ggggaagagg catttttgac tcaatcctct cagttgtact
     3901 tggagacctg cctcccagcc ctgggagatg ttttttgtat tgctcagtca taccgggcag
     3961 agcagtccag aacacgaagg cacctggctg agtacactca cgtggaagct gagtgtcctt
     4021 tcctgacttt tgacgacctc ctgaaccggt tggaggactt ggtttgtgat gtggtagatc
     4081 gaatattgaa gtcacctgca gggagcatag tgcatgagct caacccgaac tttcagcccc
     4141 ccaaacggcc tttcaaacgg atgaactatt cagatgctat cgtttggcta aaagaacatg
     4201 atgtaaagaa agaagatgga actttctatg aatttggaga agatatccca gaagctcctg
     4261 agagactgat gacagacacc attaatgaac caatcttgct gtgtcgattt cctgtggaga
     4321 tcaagtcctt ctacatgcag cgatgtcctg aggattcccg tcttactgaa tctgtcgacg
     4381 tgttgatgcc caatgttggt gagattgtgg gaggctcaat gcgtatcttt gatagtgaag
     4441 aaatactggc aggttataaa agggaaggga ttgaccccac tccctattac tggtatacgg
     4501 atcagagaaa atacggtaca tgtccccatg gaggatatgg cttgggcttg gaacgattct
     4561 taacgtggat tctgaatagg tatcacatcc gagacgtgtg cttataccct cgatttgtcc
     4621 agcgttgcac gccaagcagt aaaggtggat acggcctgaa tgatatcttt gaagcgcaga
     4681 agattgaatg gcatgaatga taaggatccg aattcgagct ccgtcgacaa gcttgcggcc
     4741 gcactcgagc accaccacca ccaccactga gatccggctg ctaacaaagc ccgaaaggaa
     4801 gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa
     4861 cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc gaatgggacg
     4921 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     4981 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     5041 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     5101 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     5161 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     5221 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     5281 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     5341 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
     5401 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgaatta
     5461 attcttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata tcaggattat
     5521 caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca ccgaggcagt
     5581 tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca acatcaatac
     5641 aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca ccatgagtga
     5701 cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc tttccagact tgttcaacag
     5761 gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg
     5821 attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta caaacaggaa
     5881 tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca cctgaatcag
     5941 gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg agtaaccatg
     6001 catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat tccgtcagcc
     6061 agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg ccatgtttca
     6121 gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca cctgattgcc
     6181 cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg gaatttaatc
     6241 gcggcctaga gcaagacgtt tcccgttgaa tatggctcat aacacccctt gtattactgt
     6301 ttatgtaagc agacagtttt attgttcatg accaaaatcc cttaacgtga gttttcgttc
     6361 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     6421 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     6481 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     6541 aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     6601 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     6661 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     6721 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     6781 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     6841 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     6901 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     6961 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     7021 ctggcctttt gctggccttt tgctcacat
//
