Team:NYMU-Taipei/Project/pH Sensor

Motivation

 * When our system arrives in the intestine (pH is higher), it senses the pH change and starts to work.
 * In this subsystem, we are going to create a pH sensor which senses high pH's.

Goal

 * The pH sensor can sense a high pH condition and start gene expression.

Regulation of nhaA gene expression

 * Na1-Induced Transcription of nhaA, Which Encodes an Na1/H1 Antiporter in Escherichia coli, Is Positively Regulated by nhaR and Affected by hns (N. DOVER etc.,1996)
 * Molecular physiology of the Na+/H+ antiporter in Escherichia coli (E Padan and S Schuldiner, 1994)



Structure of nhaA gene promoter
00000000011111111112222222222344444444455555555556666666666777777777788888888889 12345678901234567890123456789012345678901234567890123456789012345678901234567890 ACGACAAGCTGGATTATTTTTGAAATATTGGCCTAACAAGCATCGCCGACTGACAACAAATTAATTATTACTTTTCCTAA TTAATCCCTCAGGAATCCTCACCTTAAGCTATGATTATCTAGGCTTAGGGTCACTCGTGAGCGCTTACAGCCGTCAAAAA CGCATCTCACCGCTGATGGC G CAAATTCTTC AATAGCTCGTAAAAAACGAATTATTCC TA CACTATAATCTGA TTTTAA  C G ATGATT  CGTGCGGGG  TAAAAT  AG  TAAAAA  CGATCTA  T  TCACCT GA AAGAGA AATAAAAA
 * Transcription of nhaA, the Main Na+/H+ Antiporter of Escherichia coli, Is Regulated by Na+ and Growth Phase (Nir Dover and Etana Padan et al.,2001)
 * 17,189 -17,488 in E.coli K12 genome from Ecocyc (300 bp upstream of nhaA gene)


 * G is the start of NhaR binding site
 * TTTTAA is the first -35 of P1
 * ATGATT is the second -35 of P1
 * TAAAAT is the first -10 of P1
 * TAAAAA is the second -10 of P1; A in the TAAAAA is the first TSS of P1
 * AAGAGA is the S.D. (Shine-Dalgarno) sequence in RBS
 * There are 3 nhaA promoter sequences protected by NhaR from DNase I digestion
 * AATAGCTCGTAAAAAACGAATTATTCC
 * CACTATAATCTGA TTTTAA  CG  ATG
 * CGTGCGGGG TAAAAT  AG  TAAAAA  CGATCTA  T  TCACCT ; T in T  TCACCT is the second TSS of P1
 * The extracted DNA sequence should include the NhaR binding site
 * The NhaR binding site defined in (Nir Dover and Etana Padan et al.,2001) is 120 bp long
 * The PCR product derived from primers designed by Henry is 274 bp long