The argininosuccinate lyase gene of Chlamydomonas reinhardtii: an important tool for nuclear transformation and for correlating the genetic and molecular maps of the ARG7 locus.
TLDR
Using a particle gun, a reliable nuclear transformation system has been established by complementing three mutants deficient in ASL activity with the wild‐type ASL gene and analysis of the transformants reveals variable patterns of integration of the transforming DNA into the nuclear genome.Abstract:
The argininosuccinate lyase (ASL) gene of Chlamydomonas reinhardtii has been cloned using four oligonucleotide probes corresponding to highly conserved regions of the ASL polypeptide sequence. The identity of the gene was confirmed by partial sequencing. It is unique, contains several introns and spans a region less than 7.8 kb that includes highly repetitive sequences. Using a particle gun, a reliable nuclear transformation system has been established by complementing three mutants deficient in ASL activity with the wild-type ASL gene. Analysis of the transformants reveals variable patterns of integration of the transforming DNA into the nuclear genome. Previous work has mapped the mutations in the mutants arg2 and arg7 to either end of the ARG7 locus 1.0 to 1.6 recombination map units apart. Our transformation results show that these two mutations are located within a region of 7.8 kb. This allows for the first correlation of the recombination map and the molecular map at the ARG7 locus and indicates a high recombination frequency in this region of the nuclear genome.read more
Citations
More filters
Journal ArticleDOI
High-frequency nuclear transformation of Chlamydomonas reinhardtii.
TL;DR: The availability of efficient nuclear and chloroplast transformation in Chlamydomonas provides specific advantages for the study of chloropleft biogenesis, photosynthesis, and nuclear-chloroplast genome interactions.
Journal ArticleDOI
Biofuels from algae: challenges and potential.
TL;DR: This article attempts to elucidate the major challenges to economic algal biofuels at scale, and improves the focus of the scientific community to address these challenges and move algalBiofuels from promise to reality.
Journal ArticleDOI
Transgenic expression of aminoglycoside adenine transferase in the chloroplast: a selectable marker for site-directed transformation of chlamydomonas
TL;DR: Targetted gene disruption mutants of loci required for photosynthesis, tscA and psaC, were obtained and a gene disruption of an unidentified open reading frame, ORF472, remained heteroplasmic, suggesting that it has a vital function.
Journal ArticleDOI
Chlorophyll a oxygenase (CAO) is involved in chlorophyll b formation from chlorophyll a
TL;DR: It is demonstrated that a chlorophyll a oxygenase is involved inchlorophyll b formation and that an overlapping region of a nuclear genome was deleted in all mutants and that this encodes a protein whose sequence is similar to those of methyl monooxygenases.
References
More filters
Book
Molecular Cloning: A Laboratory Manual
TL;DR: Molecular Cloning has served as the foundation of technical expertise in labs worldwide for 30 years as mentioned in this paper and has been so popular, or so influential, that no other manual has been more widely used and influential.
Journal ArticleDOI
DNA sequencing with chain-terminating inhibitors
TL;DR: A new method for determining nucleotide sequences in DNA is described, which makes use of the 2',3'-dideoxy and arabinon nucleoside analogues of the normal deoxynucleoside triphosphates, which act as specific chain-terminating inhibitors of DNA polymerase.
Journal ArticleDOI
Detection of specific sequences among DNA fragments separated by gel electrophoresis.
TL;DR: This paper describes a method of transferring fragments of DNA from agarose gels to cellulose nitrate filters that can be hybridized to radioactive RNA and hybrids detected by radioautography or fluorography.
Journal ArticleDOI
A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity
TL;DR: A technique for conveniently radiolabeling DNA restriction endonuclease fragments to high specific activity is described, and these "oligolabeled" DNA fragments serve as efficient probes in filter hybridization experiments.
Journal ArticleDOI
Unidirectional digestion with exonuclease III creates targeted breakpoints for DNA sequencing.
TL;DR: A method is described for the rapid generation and cloning of deletion derivatives well-suited for the sequencing of long stretches of DNA based on two useful features of exonuclease III: processive digestion at a very uniform rate and failure to initiate digestion at DNA ends with four-base 3'-protrusions.