LOCUS       sequence-180622-        4238 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCR2.1-based plasmid holding a 309-bp fragment containing the 
            exon-1-exon-2 splice border of normal human beta-globin mRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4238)
  AUTHORS   Patsali P, Papasavva P, Christou S, Sitarou M, Antoniou MN, Lederer 
            CW, Kleanthous M
  TITLE     Relative and Absolute Quantification of Aberrant and Normal Splice 
            Variants in HBB(IVSI-110 (G > A)) beta-Thalassemia.
  JOURNAL   Int J Mol Sci. 2020 Sep 11;21(18). pii: ijms21186671. doi: 
            10.3390/ijms21186671.
  PUBMED    32933098
REFERENCE   2  (bases 1 to 4238)
  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: "Int J Mol 
            Sci."; date: "2020-09-11"; volume: "21(18). pii: ijms21186671. doi";
            pages: " 10.3390/ijms21186671"
COMMENT     Sequence Label: pCR2.1_HBB_N
FEATURES             Location/Qualifiers
     source          1..4238
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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)."
     primer_bind     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(671..690)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(672..690)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(697..714)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(697..713)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(706..728)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      854..1282
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(941..960)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1151..1172
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             1626..2420
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1680..1699)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2290..2309
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             join(2438..2506,2507..3298)
                     /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     complement(2656..2675)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3469..4057
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3958..3977
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4211..4228
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagcttg
      241 gtaccgagct cggatccact agtaacggcc gccagtgtgc tggaattcgg cttttcacta
      301 gcaacctcaa acagacacca tggtgcatct gactcctgag gagaagtctg ccgttactgc
      361 cctgtggggc aaggtgaacg tggatgaagt tggtggtgag gccctgggca ggctgctggt
      421 ggtctaccct tggacccaga ggttctttga gtcctttggg gatctgtcca ctcctgatgc
      481 tgttatgggc aaccctaagg tgaaggctca tggcaagaaa gtgctcggtg cctttagtga
      541 tggcctggct cacctggaca acctcaaggg cacctttgcc acactgagtg agctgcactg
      601 tgaagccgaa ttctgcagat atccatcaca ctggcggccg ctcgagcatg catctagagg
      661 gcccaattcg ccctatagtg agtcgtatta caattcactg gccgtcgttt tacaacgtcg
      721 tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc
      781 cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct
      841 gaatggcgaa tggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
      901 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
      961 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     1021 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     1081 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     1141 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     1201 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     1261 taacaaaaat ttaacgcgaa ttttaacaaa attcagggcg caagggctgc taaaggaagc
     1321 ggaacacgta gaaagccagt ccgcagaaac ggtgctgacc ccggatgaat gtcagctact
     1381 gggctatctg gacaagggaa aacgcaagcg caaagagaaa gcaggtagct tgcagtgggc
     1441 ttacatggcg atagctagac tgggcggttt tatggacagc aagcgaaccg gaattgccag
     1501 ctggggcgcc ctctggtaag gttgggaagc cctgcaaagt aaactggatg gctttcttgc
     1561 cgccaaggat ctgatggcgc aggggatcaa gatctgatca agagacagga tgaggatcgt
     1621 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
     1681 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
     1741 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
     1801 aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
     1861 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
     1921 ggcaggatct cctgtcatcc caccttgctc ctgccgagaa agtatccatc atggctgatg
     1981 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
     2041 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
     2101 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc
     2161 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
     2221 aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc
     2281 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
     2341 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
     2401 ttcttgacga gttcttctga attgaaaaag gaagagtatg agtattcaac atttccgtgt
     2461 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     2521 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     2581 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     2641 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     2701 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     2761 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     2821 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     2881 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     2941 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt
     3001 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     3061 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     3121 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     3181 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     3241 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     3301 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     3361 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     3421 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     3481 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     3541 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     3601 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     3661 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     3721 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     3781 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     3841 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     3901 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     3961 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     4021 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     4081 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     4141 ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac
     4201 gaccgagcgc agcgagtcag tgagcgagga agcggaag
//
