A reporter promoter assay confirmed the role of a distal promoter NOBOX binding element in enhancing expression of GDF9 gene in buffalo oocytes.
Growth differentiation factor 9 is primarily expressed in oocytes and plays a vital role in oocyte cumulus crosstalk. Earlier studies with buffalo oocytes revealed differential expression of this gene under different media stimulation conditions which, in turn, are correlated with the blastocyst yield. In this study, different germ cell specific cis elements including a NOBOX binding elements (NBE) and several E-boxes were identified at the 5' upstream region of buffalo GDF9 gene and their potential role in GDF9 expression was investigated. Transfecting oocytes with GDF9 promoter deletion constructs harbouring the NBE reporter gene revealed a 33% increase in GFP as well as the luciferase signal signifying its role in stimulating the minimal promoter activity of GDF9 in buffalo oocytes. Site directed mutation of core binding nucleotides at NBE at 1.8 kb upstream to TSS further confirmed its role for enhancing the basal transcriptional activity of GDF9 promoter in buffalo oocytes. Current work will provide important leads for understanding the role of GDF9 in oocytes competence and designing a more physiological IVF protocol in case of buffalo.
Animal Genomics Lab, ABTC, NDRI, Karnal, India. bhaskar.biotech@gmail.com
Bhaskar
B
Roy
Sandeep
S
Rajput
Sarvesh
S
Raghav
Parveen
P
Kumar
Arpana
A
Verma
Sandeep
S
Kumar
Sachinandan
S
De
Surender Lal
SL
Goswami
Tirtha Kumar
TK
Datta
D000818:Animals; D002020:Buffaloes / Q000502:physiology*; D005260:Female; D018507:Gene Expression Regulation, Developmental; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000378:metabolism; D055429:Growth Differentiation Factor 9 / Q000235:genetics / Q000502:physiology*; D018398:Homeodomain Proteins / Q000235:genetics / Q000502:physiology*; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000502:physiology*; D011401:Promoter Regions, Genetic; D014162:Transfection / Q000662:veterinary
D055429:Growth Differentiation Factor 9; D018398:Homeodomain Proteins; D049452:Green Fluorescent Proteins