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Table 3 Strains, plasmids, and primer sequences used in this study

From: Efficient production of l-lactic acid by an engineered Thermoanaerobacterium aotearoensewith broad substrate specificity

Strains

 

Source

T. aotearoense SCUT27

Wild type strain

[11]

DH5α

E. coli cloning strain, F-endA1 glnV44 thi-1 recA1 relA1 gyrA96 deoR nupG Φ80dlacZ∆M15 ∆(lacZYA-argF)U169, hsdR17(r K -m K +), λ–

Invitrogen

LA1002

As SCUT27, but ∆pta, ∆ack

This study

Plasmids

Description

Source

pBluescript II SK(+)

Standard cloning vector, f1 ori; AmpR;

Stratagene

pBlue-aph

Derived from pBluescript II SK(+), with kanamycin expression cassette

This study

pBlue-pta-aph

Derived from pBlue-aph, with partial phosphotransacetylase (pta) gene upward of kanamycin gene

This study

pPuKAd

Homologous recombination plasmid derived from pBlue-pta-aph, with partial acetate kinase (ack) gene downward of kanamycin gene

This study

Primers

Sequence 5′ → 3′) a

Application

pta-F

AACTAGGTACCAGCGCTGTACGAAATTGCCACTC

Forward primer for pta

pta-R

GTACTGAATTCCACCCATTCCTTGTGTTATAGG

Reverse primer for pta

ack-F

GAGCGGATCCGCATAGAATTAGCTCCACTGC

Forward primer for ack

ack-R

TGACTGCGGCCGCCGACGCCTCCCATAGCTG

Reverse primer for ack

Prob-F

TATTAAGACCTGCATTTCAGAT

Forward primer for hybridization probe

Prob-R

CATTTGCCTTAGCTAACCTC

Reverse primer for hybridization probe

  1. a Underlined nucleotides indicate restriction enzyme sites.