A Simple Method for DNA Extraction from Mature Date Palm Leaves: Impact of Sand Grinding and Composition of Lysis Buffer

Abstract

Molecular marker techniques have been widely used for cultivar identification of inbred date palms (Phoenix dactylifera L.; Arecaceae) and biodiversity conservation. Isolation of highly pure DNA is the prerequisite for PCR amplification and subsequent use such as DNA fingerprinting and sequencing of genes that have recently been developed for barcoding. To avoid problems related to the preservation and use of liquid nitrogen, we examined sterile sand for grinding the date palm leaves. Individual and combined effects of sodium chloride (NaCl), polyvinylpyrrolidone (PVP) and lithium chloride (LiCl) with the cetyltrimethylammonium bromide (CTAB) method for a DNA yield of sufficient purity and PCR amplification were evaluated in this study. Presence of LiCl and PVP alone or together in the lysis buffer did not significantly improve the DNA yield and purity compared with the addition of NaCl. Our study suggested that grinding of date palm leaf with sterile sand and inclusion of NaCl (1.4 M) in the lysis buffer without the costly use of liquid nitrogen, PVP and LiCl, provides a DNA yield of sufficient purity, suitable for PCR amplification.

A Comprehensive Comparison of DNA Extraction Using Fresh and Stored Bloods in Molecular Hematology Diagnostic Study

“…The values of 260/230 OD ratio for pure nucleic acid were often higher than the respective 260/280 OD ratio values 31 . We found that the ratio at 260/230 nm wasabove 2.0, which confirmedthe extracted DNA to be of good quality 30 . Lower ratios may indicate the presence of phenols, aromatic compounds, peptides and carbohydrates contaminants which absorb near 230 nm 29,32 .…”

“…In our study, the 260/280 OD ratio wasabove of 1.8 and less than 2.0; indicating the extracted DNA to be free from protein and RNA contaminations ( Table 2). A secondary measure of nucleic acid purity was based on the 260/230 OD ratio 26,30 . The values of 260/230 OD ratio for pure nucleic acid were often higher than the respective 260/280 OD ratio values 31 .…”

Preservation and maceration of amazon açai leaflet tissue to obtain genomic DNA

“…The use of liquid nitrogen to macerate leaf tissue is advantageous because it allows rapid freezing and as a result, it avoids DNA degradation (DANNER et al, 2011). However, attainment, cost, and storage are the obstacles for its use in many laboratories (IBRAHIM et al, 2010;IHASE et al, 2016), such as those located in the Amazon.…”

“…These factors limit the routine use of liquid nitrogen in many laboratories, including those located in the Amazon. Several studies have investigated alternative methods to macerate leaflets, especially in fibrous palms (IBRAHIM et al, 2010;IHASE et al, 2016). Ibrahim et al (2010) used sterilized sand with buffer to macerate mature leaves of date palm (Phoenix dactylifera).…”

“…Several studies have investigated alternative methods to macerate leaflets, especially in fibrous palms (IBRAHIM et al, 2010;IHASE et al, 2016). Ibrahim et al (2010) used sterilized sand with buffer to macerate mature leaves of date palm (Phoenix dactylifera). Ihase et al (2016) macerated leaves of oil palm (Elaeis guineensis) in a crucible with buffer.…”