LOCUS       sequence-223478-        7438 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express human LRRC33 in 293T cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7438)
  AUTHORS   Qin Y, Garrison BS, Ma W, Wang R, Jiang A, Li J, Mistry M, Bronson 
            RT, Santoro D, Franco C, Robinton DA, Stevens B, Rossi DJ, Lu C, 
            Springer TA
  TITLE     A Milieu Molecule for TGF-beta Required for Microglia Function in 
            the Nervous System.
  JOURNAL   Cell. 2018 Jun 28;174(1):156-171.e16. doi: 
            10.1016/j.cell.2018.05.027. Epub 2018 Jun 14.
  PUBMED    29909984
REFERENCE   2  (bases 1 to 7438)
  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"; doi: 
            "10.1016/j.cell.2018.05.027"; journalName: "Cell"; date: 
            "2018-06-28- 28"; volume: "174"; issue: "1"; pages: "156-171.e16"
COMMENT     Sequence Label: ET1-human LRRC33
FEATURES             Location/Qualifiers
     source          1..7438
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        111..414
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        415..618
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     569..589
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      657..666
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2652..2687
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             2697..2714
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    join(2847..3421,3422..3424,3425..3433)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3014..3031)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3241..3260
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(3434..3436,3437..3439,3440..4153)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3434..3436,3437..3439,3440..4150)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3479..3500)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3740..3759)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4087..4108
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4276..4397
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4313..4332)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4367..4386
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4404..4859)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4541..4562)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4753..4772
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4886..4990
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4992..5349
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4992..5183
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5015..5035)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5200..5335
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5384..6178
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             5384..6178
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5438..5457)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6048..6067
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6366..6385)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6410..6457
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6786..7374
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7275..7294
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttcctgcgtt atcccctgat tctgtggata accgtattac cgccatgcat tagttattaa
       61 tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg cgttacataa
      121 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata
      181 atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag
      241 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc
      301 cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta
      361 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg
      421 cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt
      481 ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca
      541 aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag
      601 gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgtcg acgtacgcca
      661 ccatggatat gagagttcct gctcaattgc tggggttgct cttgctctgg tttcctggct
      721 cgacctggag gaacagaagc ggaacagcca cagcagcctc ccaaggagtc tgcaagttgg
      781 tgggtggagc cgctgactgc cgagggcaga gcctcgcttc ggtgcccagc agcctcccgc
      841 cccacgcccg gatgctcacc ctggatgcca accctctcaa gaccctgtgg aatcactccc
      901 tccagcctta ccctctcctg gagagcctca gcctgcacag ctgccacctg gagcgcatca
      961 gccgcggcgc cttccaggag caaggtcacc tgcgcagcct ggtactgggg gacaactgcc
     1021 tctcagagaa ctacgaagag acggcagccg ccctccacgc cctgccgggc ctgcggaggc
     1081 tggacttgtc aggaaacgcc ctgacggagg acatggcagc gctcatgctc cagaacctct
     1141 cctcgctgcg gtccgtgtcc ctggcgggga acaccatcat gcggctggac gactccgtct
     1201 tcgagggcct ggagcgtctc cgggagctgg atctgcagag gaactacatc ttcgagatcg
     1261 agggcggcgc tttcgacggc ctggctgagc tgaggcacct caacctggcc ttcaacaacc
     1321 tcccctgcat cgtggacttc gggctcacgc ggctgcgggt cctcaacgtc agctacaacg
     1381 tcctggagtg gttcctcgcg accgggggag aggctgcctt cgagctggag acgctggacc
     1441 tgtctcacaa ccagctgctg ttcttcccgc tgctgcccca gtacagcaag ttgcggaccc
     1501 tcctgctgcg cgacaacaac atgggcttct accgggacct gtacaacacc tcgtcgccga
     1561 gggagatggt ggcccagttc ctcctcgtgg acggcaacgt gaccaacatc accaccgtca
     1621 gcctctggga agaattctcc tccagcgacc tcgcagatct ccgcttcctg gacatgagcc
     1681 agaaccagtt ccagtacctg ccagacggct tcctgaggaa aatgccttcc ctctcccacc
     1741 tgaacctcca ccagaattgc ctgatgacgc ttcacattcg ggagcacgag ccccccggag
     1801 cgctcaccga gctggacctg agccacaacc agctgtcgga gctgcacctg gctccggggc
     1861 tggccagctg cctgggcagc ctgcgcttgt tcaacctgag ctccaaccag ctcctgggcg
     1921 tcccccctgg cctcttcgcc aatgctagga acatcactac acttgacatg agccacaatc
     1981 agatctcact ttgtcccctg ccagctgcct cggaccgggt gggcccccct agctgtgtgg
     2041 atttcaggaa tatggcatct ttaaggagcc tgtctctgga gggctgtggc ctgggggcat
     2101 tgccagactg cccattccaa gggacctccc tgacctactt agacctctca agcaactggg
     2161 gggttctgaa tgggagcctc gccccactcc aggatgttgc ccccatgtta caggtcctgt
     2221 ctctcaggaa catgggcctc cactccagct ttatggcgtt ggacttctct gggtttggga
     2281 atctcaggga cttagatctg tcggggaatt gcttgaccac cttcccaagg tttgggggca
     2341 gcctggccct ggagaccctg gatctccgta gaaactcgct cacagccctt ccccagaagg
     2401 ctgtgtctga gcagctgtcg agaggtctgc ggaccatcta cctcagtcag aatccatatg
     2461 actgctgtgg ggtggatggc tggggggccc tgcagcatgg gcagacggtg gccgactggg
     2521 ccatggtcac ctgcaacctc tcctccaaga tcatccgcgt gacggagctg cccggaggtg
     2581 tgcctcggga ctgcaagtgg gagcggctgg acctgggcct gacgcgtgaa ctgtacttcc
     2641 aaggtccagt tgaagatcaa gttgatccaa gattgattga tggcaagggc tccggtcacc
     2701 atcaccatca ccacggcacc actggttgga gaggtggcca tgttgtggaa ggcttggctg
     2761 gtgaattgga acaattgaga gcccgtctgg agcatcaccc acaaggccaa agagaaccag
     2821 cgtaaactag tggcgcgccg gatccgcccc tctccctccc ccccccctaa cgttactggc
     2881 cgaagccgct tggaataagg ccggtgtgcg tttgtctata tgttattttc caccatattg
     2941 ccgtcttttg gcaatgtgag ggcccggaaa cctggccctg tcttcttgac gagcattcct
     3001 aggggtcttt cccctctcgc caaaggaatg caaggtctgt tgaatgtcgt gaaggaagca
     3061 gttcctctgg aagcttcttg aagacaaaca acgtctgtag cgaccctttg caggcagcgg
     3121 aaccccccac ctggcgacag gtgcctctgc ggccaaaagc cacgtgtata agatacacct
     3181 gcaaaggcgg cacaacccca gtgccacgtt gtgagttgga tagttgtgga aagagtcaaa
     3241 tggctctcct caagcgtatt caacaagggg ctgaaggatg cccagaaggt accccattgt
     3301 atgggatctg atctggggcc tcggtacaca tgctttacat gtgtttagtc gaggttaaaa
     3361 aaacgtctag gccccccgaa ccacggggac gtggttttcc tttgaaaaac acgatgataa
     3421 tatggccaca accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
     3481 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
     3541 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     3601 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
     3661 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     3721 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
     3781 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     3841 catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga
     3901 caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag
     3961 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     4021 gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg
     4081 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     4141 gctgtacaag taaagcggcc gcgactctag atcataatca gccataccac atttgtagag
     4201 gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat
     4261 gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     4321 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     4381 ctcatcaatg tatcttaagg cgtaaattgt aagcgttaat attttgttaa aattcgcgtt
     4441 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     4501 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     4561 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     4621 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     4681 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     4741 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     4801 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     4861 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     4921 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     4981 aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga
     5041 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     5101 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     5161 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     5221 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     5281 gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     5341 ttttgcaaag atcgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg
     5401 attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca
     5461 acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt
     5521 tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg
     5581 gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga
     5641 agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca
     5701 ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct
     5761 tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac
     5821 tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc
     5881 gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt
     5941 gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt
     6001 catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg
     6061 tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat
     6121 cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc
     6181 gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc
     6241 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc
     6301 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc tagggggagg
     6361 ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac ggcaataaaa
     6421 agacagaata aaacgcacgg tgttgggtcg tttgttcata aacgcggggt tcggtcccag
     6481 ggctggcact ctgtcgatac cccaccgaga ccccattggg gccaatacgc ccgcgtttct
     6541 tccttttccc caccccaccc cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc
     6601 ggggcggcag gccctgccat agcctcaggt tactcatata tactttagat tgatttaaaa
     6661 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     6721 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     6781 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     6841 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     6901 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     6961 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     7021 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     7081 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     7141 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     7201 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     7261 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     7321 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     7381 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttct
//
