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Table 1 Oligonucleotides for QRT-PCR and genetic manipulations.

From: Interference with histidyl-tRNA synthetase by a CRISPR spacer sequence as a factor in the evolution of Pelobacter carbinolicus

Primers for QRT-PCR

Name

Purpose

Location

Sequence (5' to 3')

MA0326

 

spacer #2

CCTGGTTGAGGTTAGCGTTGA

PCR of

spacer #1

   

MA0327

 

outside CRISPR

AATTCGGTGGCCAGTTGTTC

MA0328

 

sense strand

CAGGAAGCCACCAAGGAT

PCR of hisS

Pcar_1041

   

MA0329

 

antisense strand

TGGGAGCCGAGTTGATTG

MA0441

 

sense strand

CAAACTGATTGCCGTTCCTT

PCR of hisZ

GSU3307

   

MA0442

 

antisense strand

AGGCCGATGAGTTCTACGC

Primers for construction of hisS transgenic strain MA159

Name

Purpose

Sequence (5' to 3')

MA0330

 

TGACATCTCGCTGGACCGGG

PCR on 5'

side of hisS

GSU1659

   

MA0331

 

CTATGCTAGCACTAGTTTGTAATCATGAACGTACCTACTC

CTTTAATTG

MA0332

 

GTACGTTCATGATTACAAACTAGTGCTAGCATAGCAATAC

CTGCATTG

PCR on 3'

side of hisS

GSU1659

   

MA0333

 

AGTCCATTCCTCCTGTGG

MA0334

 

AAGGGATCTATCATGAGCATATCAGGCATTAAGGG

PCR of hisS

Pcar_1041

   

MA0335

 

GCGCGGCGCGACTAGTTTCCTCGTGTCTTTTCC

MA0052

 

TGCATATGGCTCTAGAATAACTTCGTATAGC

gentamicin

marker

   

MA0053

 

TCGATAAGCTTCTAGAATAACTTCGTATAATG

Oligonucleotides for construction of chimeric CRISPR expression plasmid pMA35

Name

Purpose

Sequence (5' to 3')

MA0269

 

ACATGTCACTGCCCGCTTTCCAGTC

PCR of lacI-

taclacUV5

promoter

   

MA0270

 

GCATGCGTGTGAAATTGTTATCCGC

MA0429

 

AATTCGGTTCATCCCCGCGCATGCGGGGAACACATACAT

GAGGGCAAACGCCTTTTGGCCGGCGGCGGTTCATCCCCG

CGCATGCGGGGAACACG

synthetic

CRISPR of

spacer #1

   

MA0430

 

GATCCGTGTTCCCCGCATGCGCGGGGATGAACCGCCGCC

GGCCAAAAGGCGTTTGCCCTCATGTATGTGTTCCCCGCAT

GCGCGGGGATGAACCG

MA36R

sequencing

CGACATCATAACGGTTC

  1. Note: Within the sequence of the chimeric CRISPR, a single base pair (underlined) has been duplicated in plasmid pMA35-!, at the exact centre of spacer #1.