LOCUS       sequence-357374-        5552 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli expression of class I MHC light chain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5552)
  AUTHORS   Altman lab
  TITLE     Altman lab bacterial expression plasmids
  JOURNAL   (unpublished)
REFERENCE   2  (bases 1 to 5552)
  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"; journalName: 
            "(unpublished)"
COMMENT     Sequence Label: pTCF200 Mouse beta-2 microglobulin
FEATURES             Location/Qualifiers
     source          1..5552
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     775..794
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(814..891)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1097..1116
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1200..1219
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1200..1218
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1219..1243
                     /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             1258..1280
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1612..1629
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1682..1700)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1696..1743
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1780..2235
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1867..1886)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2077..2098
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2328..3143)
                     /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(2328..3143)
                     /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     3051..3070
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3135..3154
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3265..3853
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3754..3773
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4007..4024
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4039..4181
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4107..4126)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4267..4289
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4283..4474)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5283..5552,1..813))
                     /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 cactaccgag atatccgcac caacgcgcag cccggactcg gtaatggcgc gcattgcgcc
       61 cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct cattcagcat
      121 ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt ccgctatcgg
      181 ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac gcgccgagac
      241 agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga ccagatgctc
      301 cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg gtgtctggtc
      361 agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag caatggcatc
      421 ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga gaagattgtg
      481 caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca ccacgctggc
      541 acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc
      601 cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt gttgtgccac
      661 gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc gcgttttcgc
      721 agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga caccggcata
      781 ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt gactctcttc
      841 cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt ccgggatctc
      901 gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt
      961 tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac agtcccccgg
     1021 ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag
     1081 cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc
     1141 cggtgatgcc ggccacgatg cgtccggcgt agaggatcga gatctcgatc ccgcgaaatt
     1201 aatacgactc actatagggg aattgtgagc ggataacaat tcccctctag aaataatttt
     1261 gtttaacttt aagaaggaga tatacatatg atccagaaaa cccctcagat tcaggtttat
     1321 tcgcgtcacc cgccggagaa tggcaaacca aacatcttga actgctatgt cacgcagttc
     1381 cacccgccgc atattgagat tcaaatgtta aagaatggca aaaagatccc aaaagtcgaa
     1441 atgtcagata tgagctttag caaggattgg tcattttata tcctcgccca tactgaattt
     1501 acaccaacgg agaccgatac ctatgcctgc cgtgtgaaac acgcgtctat ggcggaaccg
     1561 aagaccgttt attgggatcg tgacatgtaa aagcttgcgg ccgcactcga gcaccaccac
     1621 caccaccact gagatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc
     1681 accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt gaggggtttt
     1741 ttgctgaaag gaggaactat atccggattg gcgaatggga cgcgccctgt agcggcgcat
     1801 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
     1861 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
     1921 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
     1981 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt
     2041 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa
     2101 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg
     2161 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat
     2221 taacgtttac aatttcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     2281 tatttttcta aatacattca aatatgtatc cgctcatgaa ttaattctta gaaaaactca
     2341 tcgagcatca aatgaaactg caatttattc atatcaggat tatcaatacc atatttttga
     2401 aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga
     2461 tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc
     2521 tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag
     2581 aatggcaaaa gtttatgcat ttctttccag acttgttcaa caggccagcc attacgctcg
     2641 tcatcaaaat cactcgcatc aaccaaaccg ttattcattc gtgattgcgc ctgagcgaga
     2701 cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc
     2761 aggaacactg ccagcgcatc aacaatattt tcacctgaat caggatattc ttctaatacc
     2821 tggaatgctg ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg
     2881 ataaaatgct tgatggtcgg aagaggcata aattccgtca gccagtttag tctgaccatc
     2941 tcatctgtaa catcattggc aacgctacct ttgccatgtt tcagaaacaa ctctggcgca
     3001 tcgggcttcc catacaatcg atagattgtc gcacctgatt gcccgacatt atcgcgagcc
     3061 catttatacc catataaatc agcatccatg ttggaattta atcgcggcct agagcaagac
     3121 gtttcccgtt gaatatggct cataacaccc cttgtattac tgtttatgta agcagacagt
     3181 tttattgttc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     3241 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     3301 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     3361 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt
     3421 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     3481 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     3541 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     3601 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     3661 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     3721 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     3781 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     3841 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     3901 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     3961 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     4021 cgaggaagcg gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc
     4081 acaccgcata tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     4141 tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac
     4201 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     4261 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc
     4321 agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat
     4381 ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg
     4441 ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc
     4501 tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg
     4561 atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc
     4621 ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag
     4681 gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg
     4741 gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg
     4801 ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg
     4861 attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga
     4921 gcacgatcat gcgcacccgt ggggccgcca tgccggcgat aatggcctgc ttctcgccga
     4981 aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag attccgaata
     5041 ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga
     5101 cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca gtcataagtg
     5161 cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg aaggctctca
     5221 agggcatcgg tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc
     5281 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     5341 aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt
     5401 gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg
     5461 tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata
     5521 taacatgagc tgtcttcggt atcgtcgtat cc
//
