Nucleotide sequences of the continuous and separated petA, petB and petD chloroplast genes in Chlamydomonas reinhardtii

Nucleotide sequences of the continuous and separated petA, petB and petD chloroplast genes in Chlamydomonas reinhardtii

            S. Büschlen, Y. Choquet, R. Kuras, F.-A. Wollman

            Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, 75005 Paris, France

Chlamydomonas reinhardtii is a unicellular green alga, and has been used in studying genetic expression in eucaryotic cells by oxygenic photosynthesis. Using probes from the corresponding pet genes in spinach, the petA, petB and petD genes were mapped on the chloroplast genome of C. reinhardtii. The experiment revealed that petB was located on the chloroplast DNA restriction fragments Ba11 and R20, numbered after Rochaix. Where as petA and petD genes both stayed on the same DNA restriction fragments Ba9 and R26. This proved that the petB and petD genes are not located on the same chloroplast genome and are separate from psbB-psbF-petB-petD cluster.

Reference

FEBS Letters, Volume 284, Issue 2, 24 June 1991, Pages 257-262
S. Büschlen, Y. Choquet, R. Kuras, F.-A. Wollman

http://www.sciencedirect.com/science/article/pii/0014579391806983

Clean-up of Successfully Amplified petB From DNA 4-1 through 4-9

Brandon Jones and Duong V. performed a PCR cleanup, seven out of the nine DNA’s were cleaned successfully. Two of the samples were poorly amplified (4-5 and 4-8) and were not able to be used in the PCR cleanup results as illustrated in Table 1. The PCR results in Table 1 shows moderate to high yield for samples: 4-1, 4-3  and a high yield for samples: 4-2, 4-4, 4-6, 4-7 and, 4-8.  The cleanup process was performed by using Exonuclease I and Alkaline Phosphatase combined with PCR mixture for further purification.Our results shown in Table 1 below are from using the nano-drop spectrophotometer.

Table 1: Quantification of the enzyme treated PCR products

Sample ID	ng/ul
4-1B	389.01
4-2B	496.81
4-3B	366.41
4-4B	463.28
4-6B	557.95
4-7B	521.77
4-9B	545.84

petB intron for 4-1 through 4-9

Brandon J. and Duong V. used PCR to amplify the petB region of  nine DNAs. A total of nine samples were used including a marker lane 1, and water lane 11, as illustrated in the photo (Figure 1). The nine samples of extracted DNA were then prepped for gel electrophoresis. The gel was created, then it was placed in the electrophoresis equipment, samples were added to it, and the electrophoresis equipment was turned on to create a charge for a period of time. This was all done followed by instructions given from Dr. Jarrell.  Based on the Molecular Mass Ruler these products  are larger than 1000 bp because the products in Figure 1 ran slower than  the Mass Ruler sample provided.

The results of the gel electrophoresis is listed in the table below (Table 1). The fluorescent bands contained near/in the wells is unknown, it could be caused by some of the DNA stain coming out of solution, causing it to stay in the wells or poorly migrate out of them. Also alternative explanation is that the running buffer had been previously used and some DNA stain was left over from prior runs.

Figure 1

PCR Success Results:

++ High

+ Low

-No Success

Table 1

Lane	DNA code	Genus Species	PCR Results
1	Blank	Gel Marker	N/A
2	4-1	Chiloschista segawae	++
3	4-2	Phalaenopsis viridis	++
4	4-3	Rhynchostylis gigantea	+
5	4-4	Holcoglossum subulifolium	+
6	4-5	Phalaenopsis equestris	–
7	4-6	Rhynchostylis coelestis	+
8	4-7	Angraecum didieri	++
9	4-8	Aerangis luteo-alba	–
10	4-9	Cleisostoma schneideri	++
11	Negative control	Water	N/A

Grosourdya appendiculata

Grosourdya appendiculata Rchb.f. is part of the Kingdom Plantae, Phylum Tracheophyta, Class Liliopsida, Order Asparagales, and Family Orchidaceae. This genus was first described by Carl Blume in 1825. Grosourdya is a small size growing specie. In 1867 Reichenbach established it as a monotypic genus, meaning it has only one  species. Reichenbach provided its current accepted name, and in 1972 Garay added ten other species to Grosourdya.

Grosourdya appediculata Photo taken by Dr. Jarrell

Grosourdya appendiculata is located from the Andamans to South Vietnam, through Thailand, Malaya to Java, and Borneo to the Philippines. Grosourdya appendiculata is normally distributed at low elevations near sea level. This species has also been found near Merek in North Sumatra at 1200m by J.J. Smith in 1933.

Grosourdya appendiculata Size:

• Short-stemmed epiphyte producing about 6-8 leaves.
• Leaves are about 14X2cm
• Flowers produced, only last about one day and its opening is about 0.75-1.5cm broad.

Grosourdya appendiculata provided by Jarrell, Photographed by Brandon J.

Source:

*J.B Comber. Orchids of Sumatra (2001), S. 819-820

*J.B. Comber. Orchids of Java (1990), S. 303

*Seidenfaden, G. in Opara Botanica 95 (1988), S. 319-324