Tandem duplicate expression patterns are conserved between maize haplotypes of the α‐zein gene family

Abstract

Tandem duplication gives rise to copy number variation and subsequent functional novelty among genes as well as diversity between individuals in a species. Functional novelty can result from either divergence in coding sequence or divergence in patterns of gene transcriptional regulation. Here, we investigate conservation and divergence of both gene sequence and gene regulation between the copies of the α-zein gene family in maize inbreds B73 and W22. We used RNA-seq data generated from developing, self-pollinated kernels at three developmental stages timed to coincide with early and peak zein expression. The reference genome annotations for B73 and W22 were modified to ensure accurate inclusion of their respective α-zein gene models to accurately assess copy-specific expression. Expression analysis indicated that although the total expression of α-zeins is higher in W22, the pattern of expression in both lines is conserved. Additional analysis of publicly available RNA-seq data from a diverse population of maize inbreds also demonstrates variation in absolute expression, but conservation of expression patterns across a wide range of maize genotypes and α-zein haplotypes.

Editing the 19 kDa alpha‐zein gene family generates non‐opaque2‐based quality protein maize

“…As the alpha‐zein gene family is made up of many gene copies partitioned into separate subfamilies, it was important to identify which copies are expressed and to what degree. Previously, an RNAseq study compared zein transcription between two inbred lines with differing alpha‐zein haplotypes, B73 and W22 (Hurst et al ., 2021 ). The results were used to prioritize the specific gene copies which dominate the expression profile, in order to ensure complete knock‐out and avoid residual expression compromising the hypothesized phenotype.…”

“…The results were used to prioritize the specific gene copies which dominate the expression profile, in order to ensure complete knock‐out and avoid residual expression compromising the hypothesized phenotype. Of note, the oldest family of 19 kDa gene copies, the D family on chromosome 1, were determined to be pseudogenes and not considered viable target genes (Dong et al ., 2016 ; Hurst et al ., 2021 ). All putative target sites, defined as a 20 bp string with a 5′‐NGG‐3′ adjacent motif, were compared in a pair‐wise fashion.…”

“…A custom Python script was used to extract all sequences with a 5′‐NGG‐3′ PAM sequence and make pairwise comparisons between them. A transcriptomic study was conducted to prioritize gene copies by editing potential importance (Hurst et al ., 2021 ). Seed sequence match, location of mismatches and off‐target potential were considered.…”