LOCUS       sequence-308672-        5431 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Recombination-Induced Tag Exchange (RITE) Cassette, HA-6xHIS to a V5
            epitope tag, Hygro selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5431)
  AUTHORS   Terweij M, van Welsem T, van Deventer S, Verzijlbergen KF, 
            Menendez-Benito V, Ontoso D, San-Segundo P, Neefjes J, van Leeuwen F
  TITLE     Recombination-induced tag exchange (RITE) cassette series to monitor
            protein dynamics in Saccharomyces cerevisiae.
  JOURNAL   G3 (Bethesda). 2013 Aug 7;3(8):1261-72. doi: 10.1534/g3.113.006213.
  PUBMED    23708297
REFERENCE   2  (bases 1 to 5431)
  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: "10.1534/g3.113"; 
            journalName: "G3 (Bethesda)"; date: "2013-08-7- 7"; volume: "3"; 
            issue: "8"; pages: "1261-72"
COMMENT     Sequence Label: pMT001
FEATURES             Location/Qualifiers
     source          1..5431
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(237..692)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(374..395)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     586..605
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     818..840
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     832..849
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     833..849
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     856..875
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        856..874
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    928..961
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             970..996
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             997..1014
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      1035..1222
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1090..1114)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            complement(1341..2915)
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     terminator      complement(1341..1537)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(1543..2571)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             complement(1543..2571)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     promoter        complement(2572..2915)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     protein_bind    3010..3043
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3108..3164
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     promoter        complement(3171..3189)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3210..3226)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3210..3226)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3223..3245)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3234..3250
                     /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(join(3258..3264,3265..3282,3283..3288))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3303..3324
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3441..3458)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3612..4200)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3692..3711)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4371..5162,5163..5231))
                     /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     4994..5013
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5232..5336)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5404..5422
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggaaat
      241 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
      301 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
      361 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
      421 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
      481 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
      541 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
      601 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
      661 cgccgcgctt aatgcgccgc tacagggcgc gtccattcgc cattcaggct gcgcaactgt
      721 tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt
      781 gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg
      841 acggccagtg aattgtaata cgactcacta tagggcgaat tgggcccaat gcattggcgc
      901 cgcggccgcg gtggatctgg tggatctata acttcgtata atgtatgcta tacgaagtta
      961 tcaggtacct acccatacga tgttcctgac tatgcgcacc atcatcacca tcacactagt
     1021 taacttctaa ataagcgaat ttcttatgat ttatgatttt tattattaaa taagttataa
     1081 aaaaaataag tgtatacaaa ttttaaagtg actcttaggt tttaaaacga aaattcttat
     1141 tcttgagtaa ctctttcctg taggtcaggt tgctttctca ggtatagtat gaggtcgctc
     1201 ttattgacca cacctctacc ggcagatccg ctagggataa cagggtaagg atccgcattg
     1261 tactgagagt gcaccataat catcgatgaa ttcgagctcg ttttcgacac tggatggcgg
     1321 cgttagtatc gaatcgacag cagtatagcg accagcattc acatacgatt gacgcatgat
     1381 attactttct gcgcacttaa cttcgcatct gggcagatga tgtcgaggcg aaaaaaatat
     1441 aaatcacgct aacatttgat taaaatagaa caactacaat ataaaaaaac tatacaaatg
     1501 acaagttctt gaaaacaaga atctttttat tgtcagtact gattattcct ttgccctcgg
     1561 acgagtgctg gggcgtcggt ttccactatc ggcgagtact tctacacagc catcggtcca
     1621 gacggccgcg cttctgcggg cgatttgtgt acgcccgaca gtcccggctc cggatcggac
     1681 gattgcgtcg catcgaccct gcgcccaagc tgcatcatcg aaattgccgt caaccaagct
     1741 ctgatagagt tggtcaagac caatgcggag catatacgcc cggagccgcg gcgatcctgc
     1801 aagctccgga tgcctccgct cgaagtagcg cgtctgctgc tccatacaag ccaaccacgg
     1861 cctccagaag aagatgttgg cgacctcgta ttgggaatcc ccgaacatcg cctcgctcca
     1921 gtcaatgacc gctgttatgc ggccattgtc cgtcaggaca ttgttggagc cgaaatccgc
     1981 gtgcacgagg tgccggactt cggggcagtc ctcggcccaa agcatcagct catcgagagc
     2041 ctgcgcgacg gacgcactga cggtgtcgtc catcacagtt tgccagtgat acacatgggg
     2101 atcagcaatc gcgcatatga aatcacgcca tgtagtgtat tgaccgattc cttgcggtcc
     2161 gaatgggccg aacccgctcg tctggctaag atcggccgca gcgatcgcat ccatggcctc
     2221 cgcgaccggc tgcagaacag cgggcagttc ggtttcaggc aggtcttgca acgtgacacc
     2281 ctgtgcacgg cgggagatgc aataggtcag gctctcgctg aattccccaa tgtcaagcac
     2341 ttccggaatc gggagcgcgg ccgatgcaaa gtgccgataa acataacgat ctttgtagaa
     2401 accatcggcg cagctattta cccgcaggac atatccacgc cctcctacat cgaagctgaa
     2461 agcacgagat tcttcgccct ccgagagctg catcaggtcg gagacgctgt cgaacttttc
     2521 gatcagaaac ttctcgacag acgtcgcggt gagttcaggc tttttaccca tggttgttta
     2581 tgttcggatg tgatgtgaga actgtatcct agcaagattt taaaaggaag tatatgaaag
     2641 aagaacctca gtggcaaatc ctaacctttt atatttctct acaggggcgc ggcgtgggga
     2701 caattcaacg cgtctgtgag gggagcgttt ccctgctcgc aggtccgcag cgaggagccg
     2761 taatttttgc ttcgcgccgt gcggccatca aaatgtatgg atgcaaatga ttatacatgg
     2821 ggatgtatgg gctaaatgta cgggcgacag tcacatcatg cccctgagct gcgcacgtca
     2881 agactgtcaa ggagggtatt ctgggcctcc atgtcgctgg ccgggtgacc cggcggggac
     2941 gaggcaagct aaacagatct ggcgcgcctt aattaaccct atgcggtgtg aaataccgca
     3001 cagtctagaa taacttcgta taatgtatgc tatacgaagt tatcagtcga cggtaagcct
     3061 atccctaacc ctctcctcgg tctcgattct tgtacataag cggccgcgaa ttcgagctcg
     3121 gtacccgggg atcctctaga gtcgacctgc aggcatgcaa gcttttgttc cctttagtga
     3181 gggttaattt cgagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     3241 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     3301 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     3361 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     3421 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     3481 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     3541 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     3601 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     3661 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     3721 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     3781 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     3841 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     3901 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     3961 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     4021 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     4081 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     4141 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     4201 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     4261 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     4321 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     4381 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     4441 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     4501 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     4561 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     4621 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     4681 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     4741 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     4801 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     4861 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     4921 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     4981 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     5041 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     5101 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     5161 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     5221 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     5281 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     5341 tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat
     5401 aaaaataggc gtatcacgag gccctttcgt c
//
