Category Archives: ycf1

Primer development for the plastid region ycf1 in Annonaceae and other magnoliids

This article identifies the usability of ycf1 using primers to identify the plasid gene within the magnoliid taxa. This hypothetical chloroplast open reading frame 1, more commonly identified as ycf1, codes a region of 5,500 base pairs, and is virtually present in many plastid genomes. It is typically used  in analyzing orchids and Pinus, making the use of it in magnoliids unique and somewhat rarely studied. Using an array of species across many families, the samples were able to specifically identify the usage of ycf1. An array of primers were used to amplify various portions, to amplify various nucleotide regions. By sequencing roughly 2000 base pairs within ycf1 in over 20 individuals, the researchers were able to examine the phylogenetic utility relative to other plastid regions. The genera Deeringothamnusand Asimina (Annonaceae) was used due to its close proximity. Through various PCR deviations using different primers revealed the discrepancy of ycf1 among species, discovering the higher number of informative characters. Finally, the resulting factors proved that ycf1 proves to be easily amplified and sequenced vastly more than other common markers such as matK and trnL-F. The study indicates that ycf1 has great potential utility.

 

References:

Neubig, K., & Abbott, J. (2010). Primer development for the plastid region ycf1 in annonaceae and other magnoliids. American Journal of Botany, Retrieved from http://www.amjbot.org/content/97/6/e52.full

Highly variable YCF1 useful for evaluating plant phylogeny at low taxonomic levels.

Twelve genera scanned to find highly variable regions of the chloroplast genome.Twenty-three loci were selected, the most variable were intergenic regions ycf1-a, trnK, rpl32-trnL, and trnH-psbA, followed by trnSUGA-trnGUCC, petA-psbJ, rps16-trnQ, ndhC-trnV, ycf1-b, ndhF, rpoB-trnC, psbE-petL, and rbcL-accD. Three loci, trnSUGA-trnGUCC, trnT-psbD, and trnW-psaJ, showed very high nucleotide diversity. The chloroplast genome is often used to determine plant phylogeny but becomes difficult in closely related species. Finding regions with more variability allows lower level taxonomy. The ycf1 region was found to have the most variability among the regions looked at. To solve phylogenetic problems at the species level,regions with very high evolutionary rates are needed. Greater availability of such regions will increase the ability to resolve such identification problems.

Reference:

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3325284/?tool=pmcentrez

Wenpan Dong,1,2 Jing Liu,1,3 Jing Yu,1,3 Ling Wang,2 and Shiliang Zhou1*

Cadmium Tolerance Mediated by Yeast AP-1 Protein requires the Presence of Ycf1

In the article Cadmium Tolerance Mediated by the Yeast AP-1 Protein Requires the Presence of an ATP-binding Casette Transporter-encoding gene, Ycf1, elevations in gene dosage of the protein yAP-1 in Saccharomyces cereuisiae showed an increase in the tolerance of drugs that contain the toxic heavy metal cadmium. The increase in tolerance occurs because of an overproduction of yAP-1. The ycf1 gene is required for for yAP-1 to have a normal tolerance of cadmium. Mutant strains of yeast that do not have the ycf1 gene are hypersensitive to the effects of cadmium because of the over expression of yAP-1. This data established that an important physiological target of yAP-1 transcriptional regulation is the ycf1 gene.

References:

John A. WemmieS,  Mark S. SzczypkaO, Dennis J. ThieleOn, and W. Scott Moye-RowleySII. (1994).Cadmium  Tolerance  Mediated  by

the Yeast AP-1 Protein  Requires the  Presence of an  ATP-binding  Cassette  Transporter-encoding Gene, YCFl*. The Journal of Biological Chemistry. 269, 32592-32597


Phylogenetic Utility of ycf1 in Orchids: A Plastid Gene More Variable Than matK

This article discussed how the researchers used the ycf1 region of a plastid protein compared to the matk region to determine which would give them more useful information when at species level of the Orchidaceae. They used wild and cultivated orchids for the study. After extracting the DNA of the orchids, they attempted to amplify the 3 prime end using both ycf1 and matK. They determined that while matK is the most commonly used, ycf1 returned a better result on the species level. Ycf1 had a greater variability level than that of matK. When compared to ITS, ycf1 was able to give a longer phylogenetic tree that either of the previous two mentioned. They tested ycf1, matK, and ITS with species level and tribe level. ITS was more useful and returned better variability results when it came to the species Elleanthus and Sobralia, but ycf1 preformed better at the tribe level when tested on the tribe Vandeae. They mentioned that minor incongruences  may have played a role in the results they received, but that it was common for this to occur when severl data sets were compared together. While ycf1 gives the best results when used with orchids, they mentioned that further research could also be done to determine if ycf1 would be just as useful when used with other plant taxa.

References:

Neubig, K. M., Whitten, W., Carlsward, B. S., Blanco, M. A., Endara, L., Williams, N. H., & Moore, M. (2009). Phylogenetic utility of ycf1 in orchids: a plastid gene more variable than matK. Plant Systematics & Evolution, 277(1/2), 75-84. doi:10.1007/s00606-008-0105-0

Use of the chloroplast gene ycf1 for the genetic differentiation of pine nuts obtained from consumers experiencing dysgeusia.

Pine nuts are part of traditional cooking in many places of the world, including United States over the past ten years. However, over the past three years, the U.S. Food and Drug Administration (US FDA) field received 411 complaints from the consumers regarding the taste disturbance, known as dysguesia, of pine nuts. By using gas chromatography with flame ionization detection to analyze the fatty acid, previous study implicated nuts from Pinus armandii as the causative species for similar taste disturbances. Unfortunately, this method was found to provide insufficient species resolution to link consumer complaint samples to a single species of pine, particularly when samples contained mixture of pine nuts. Therefore, in this study, a DNA based method was used for differentiating pine nut samples using ycf1 chloroplast gene. Although, the exact reason associated dysguesia is still not known, researchers found that 15 of 15 samples from consumer complaints contained at least some Pinus armandii, confirming the association of this species with taste disturbances.

Reference

Handy, Sara, Matthew  LastParks,  et al. “Use of the chloroplast gene ycf1 for the genetic differentiation of pine nuts obtained from consumers experiencing dysgeusia.” J. Agric. Food Chem. 59.20 (2011): 10995–11002. Web. 5 May. 2012. <http://pubs.acs.org/doi/full/10.1021/jf203215v>.

Insertion or Deletion Mutations; matK, ycf1 and ycf2

A study was done to determine the gene responsible for chlorophyll deficiency in Wogon-Sugi. Wogon-Sugi is a mutant that is inherited within the cytoplasm of Cryptomeria japonica. Chloroplast DNA sequencing was done on 16 wild-type plants and 16 Wogon-Sugi plants. Yaka-Sugi (wild-type) was used as the maternal plant and Wogon-Sugi was used as the paternal plant during an artificial controlled cross. After growing the seeds for a year, they were examined by a chloroplast DNA marker, based off of Wogon-Sugi.

Ycf1, ycf2 and matK were targeted as candidates for the mutation that is responsible for the phenotype Wogon-Sugi. In Wogon-Sugi plants, an insertion mutation of 19-bp was found in the matK coding region. Because of this insertion, the matK reading frame was disrupted. Ycf1 was repeated on 66-bp and ycf2 was repeated on 33-bp.  These repeats were found in both Wogon-Sugi and the wild-types, Yaka-Sugi. The ycf1 and ycf2 mutations did not shift the reading frames. It was concluded that the matK insertional sequence is the mutant specific to Wogon-Sugi chloroplast genome and is also responsible for the chlorophyll deficiency.

References

Tomonori Hirao , Atsushi Watanabe , Manabu Kurita,  Teiji Kondo, Katsuhiko Takata. “A frameshift mutation of the chloroplast matK coding region is associated with chlorophyll deficiency in the Cryptomeria japonica Virescent mutant Wogon-Sugi.” Curr Genet (2009) 55:311–321. http://web.ebscohost.com/ehost/detail?vid=3&hid=11&sid=1e411bd2-1e45-472a-8f9a-df40aacfa47440sessionmgr4&bdata=JnNpdGU9ZWhvc3QtbGl2ZSZzY29wZT1zaXRl#db=a9h&AN=41041717

“Functional Comparison between YCF1 and MRP1 Expressed in Sf21 Insect Cells”

In the article “Functional Comparison between YCF1 and MRP1…” the function of YCF1 in animal and yeast cells were examined and YCF1 and MRP1 expressed in insect cells. The authors found some similarity in the ATP-dependent transport in YCF1 and yeast. They also found that YCF1 and MRP1 have a significant similarity in extensive amino acid sequence.The functions of YCF1 in animal and yeast cells were extensively studied in the insect cells of Sf21. The authors concluded that they do have some relationship in the localizing of the protein. They also concluded that the functions of YCF1 and MRP1 are similar but are not identical.

 

 

Xiao-Qin Rena, Tatsuhiko Furukawaa, 1, Zhe-Sheng Chena, Hiroshi Okumuraa, Shunji Aokib, Tomoyuki Sumizawaa, Ayako Tania, Masaharu Komatsua, Xiao-Dong Meia, Shin-ichi Akiyama. 25 February 2000. Functional Comparison between YCF1 and MRP1 Expressed in Sf21 Insect Cells. Biochemical and Biophysical Research Communications Volume: 207, Issue 2 pages 608-615.

ycf1 and ycf1 as essential genes.

The article Large Plastid Reading Frames as Essential Genes concerns the knock out of ycf1 and ycf2 to determine the function of the gene.  Through plastid transformation (process of taking foreign DNA into plasmid) and cloning of the ycf1 and ycf2 via E. coli grown on selective streptomycin plates.  It was found that knock out of either ycf1 or ycf2 results in organism failure.  Attempting to create mutants by adding an accD marker caused organism failure leading to the determination that ycf1 and ycf2 is a gene required for the organism to function.  This determination was concluded by inserting the plasmid DNA into the chloroplast.  Speculation of function of ycf1 and ycf2 suggests that the genes are required for functional structural or pathway proteins.

References:

Drescher, A., Ruf S., Calsa T., Carrer, H., Bock, R.  (2000).  The two largest chloroplast genome-encoded open reading frames of higher plants are essential genes.  The Plant Journal 22(2), 97 – 104.  York, UK: Blackwell Science Ltd.

PCR Amplification of ycf1-3720F, ycf1-5500R from 4-12; 4-23

 

 

Result:  Our marker is in the gel but in the process of making the gel the EZ-Vision stain was not added to the marker which is the reason as to why there is no ladder seen toward the left on the gel.  Only one sample failed 4-14.  The others appear to have strong amplification.  When reading the table please note that (++) denotes a strong amplificaion, amplification (+), and (-) denotes weak ampflication.  The stain is Easy Vision One 6x and stains the marker so that a ladder may be seen. Some of the amplifications also yield other amplified genes other than the target sequence.  Trevor Rawlinson and Danny Cannon conducted the PCR with the ycf1 primers.
Channel 1 (top) Results Species
Marker (-) N/A
Control (Water) (-) N/A
4-12 (++) Ceratocentron fesselii
4-13 (++) Thrixspermum bromeliforme
4-14 (-) Holcoglossum flavescens
4-15 (++) Renanthera storei
4-16 (++) Trichoglottis amesiana
4-17 (++) Omoea philippinensis
4-18 (++) Gastrochilus acutifolius
4-19 (++) Trichoglottis bipunctata
4-20 (++) Malleola constricta
4-21 (++) Phalaenopsis lobbii
Channel 2 (bottom) Results Sepcies
Marker (-) N/A
4-22 (++) Armordorum siamense